WO2023278606A1

WO2023278606A1 - Methods of quantification and identification of molecules

Info

Publication number: WO2023278606A1
Application number: PCT/US2022/035560
Authority: WO
Inventors: Ashwin Gopinath; Paul ROTHEMUND; Shane BOWEN
Original assignee: Somalogic Operating Co., Inc.
Priority date: 2021-07-01
Filing date: 2022-06-29
Publication date: 2023-01-05

Abstract

Provided herein are structures and methods for identifying one or more analyte molecules using one or more supramolecular structures. In some embodiments, identifying an analyte molecule comprises correlating a specific combination of affinity binders that are configured to form a linkage with an analyte molecule. In some embodiments, the affinity binders are provided with binding molecules comprising barcodes corresponding to the affinity binders. In some embodiments, the affinity binders are provided with reporter molecules comprising signaling molecules. In some embodiments, a nucleic acid having a sequence corresponding to the affinity binders is linked to the supramolecular structure through interactions with an analyte molecule. In some embodiments, a supramolecular structure is contacted with a reporter molecule for one or more iterations, wherein each iteration contains a different reporter molecule from the others, and wherein the specific combination of reporter molecules detected as interacting with an analyte molecule identifies said analyte molecule.

Description

METHODS OF QUANTIFICATION AND IDENTIFICATION OF MOLECULES

CROSS REFERENCE

[0001] This application claims the benefits of U.S. Provisional Application No. 63/217,706 filed July 1, 2021 which is incorporated herein by reference in the entirety for all purposes.

BACKGROUND

[0002] The concentration of proteins, the interaction between the proteins (protein-protein interactions or PPI), as well as the interaction between proteins and small molecules, are intricately linked to the health of different organs, homeostatic regulatory mechanism as well as the interaction of these systems with the external environment. Hence, quantitative information about proteins as well as PPIs is vital to create a complete picture of an individual’s health at a given time point as well as to predict any emerging health issues. For instance, the amount of stress experienced by cardiac muscles (e.g. during a heart attack) can be inferred by measuring the concentration of troponin I/II and myosin light chain present within peripheral blood. Similar protein biomarkers have also been identified, validated and are deployed for a wide variety of organ dysfunctions (e.g. liver disease and thyroid disorders), specific cancers (e.g. colorectal or prostate cancer), and infectious diseases (e.g. HIV and Zika). The interaction between these proteins are also essential for drug development and are increasingly becoming a highly sought-after dataset. The ability to identify and quantify proteins and other molecules, within a given sample of bodily fluids, is an integral component of such healthcare development.

SUMMARY

[0003] The present disclosure generally relates to structures and processes for identification and quantification of analyte molecules in a sample.

[0004] Disclosed herein, in some embodiments, is a nucleic acid having a sequence comprising a set of segments comprising a first segment and a second segment, wherein the first segment identifies a first affinity binder to an analyte molecule and the second segment identifies a second affinity binder to the analyte molecule, wherein the first affinity binder and the second affinity binder are different; wherein each of the first affinity binder and the second affinity binder is configured to bind to the analyte molecule with a binding affinity of a dissociation constant (Kd) value, less than about 1 pM, and wherein the set of segments identifies the analyte molecule. [0005] Disclosed herein, in some embodiments, is a nucleic acid having a sequence comprising a set of segments having N segments, wherein N is greater than 1, wherein each segment identifies an affinity binder in a set of affinity binders, wherein each affinity binder in the set of affinity binders is different from other affinity binders in the set of affinity binders, and wherein each affinity binder is configured to bind to an analyte molecule with a binding affinity of a dissociation constant (Kd) value, less than about 1 pM, and wherein the set of segments identifies the analyte molecule. In some embodiments, the nucleic acid of claim 2, wherein N is 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, the nucleic acid of claim 2 wherein N is greater than 10. In some embodiments, a structure comprising the nucleic acid of claim 2 linked to a supramolecular structure.

[0006] In some embodiments, for a nucleic acid described herein, the nucleic acid is formed by a process comprising: providing a supramolecular structure comprising: a core structure comprising a plurality of core molecules, a first reactive group linked to the core structure at a first location and configured to form a linkage with the analyte molecule, and a second reactive group linked to the core structure at a second location and configured to form a linkage with a first binding molecule comprising i) a first affinity binder and ii) a first tail section comprising a first barcode corresponding to the first segment of the nucleic acid; binding the analyte molecule to the first reactive group; linking i) the first affinity binder to the analyte molecule, and ii) the first binding molecule to the second reactive group via the first tail section; separating i) the first affinity binder from the analyte molecule, and ii) a portion of the first tail section from the first barcode, such that the first barcode remains linked to the second reactive group; contacting the analyte molecule with a second binding molecule comprising i) a second affinity binder and ii) a second tail section comprising a second barcode corresponding to the second segment, such that i) the second affinity binder links with the analyte molecule, and ii) the second binding molecule links to the first barcode via the second tail section; separating i) the second affinity binder from the analyte molecule, and ii) a portion of the second tail section from the second barcode, such that the second barcode remains linked to the first barcode, wherein the set of segments comprise the first barcode and the second barcode.

[0007] In some embodiments, the process for forming a nucleic acid described herein further comprises repeating steps (e)-(f) one or more times each with a different binding molecule, such that the set of segments comprises the first barcode, the second barcode, and at least one barcode corresponding to a binding molecule from the one or more times where steps (e)-(f) was repeated. In some embodiments, the sequence comprises a unique nucleic acid strand identified via DNA sequencing. In some embodiments, the process further comprises quantifying the concentration of the analyte molecule in the sample. In some embodiments, the process further comprises identifying the detected analyte molecule. In some embodiments, the analyte molecule binds with the first reactive group via a chemical bond. In some embodiments, the chemical bond comprises a covalent bond. In some embodiments, the chemical bond comprises a non-covalent bond. In some embodiments, the second reactive group comprises a single stranded nucleic acid of a repeating specific sequence. In some embodiments, the tail section of the first binding molecule comprises a double stranded nucleic acid comprising a first strand of a first barcode sequence and a second strand of a sequence complementary to a repeating sequence of the second reactive group. In some embodiments, the tail section of the binding molecule links with the second reactive group via hybridization of nucleic acids. In some embodiments, the tail section of the binding molecule binds to the second reactive group via formation of a phosphodiester bond between nucleic acids. In some embodiments, the tail section of each binding molecule of the one or more binding molecules further comprises one or more reactive binders configured to link the corresponding affinity binder to the corresponding barcode. In some embodiments, the process, prior to step (c), further comprises incubating a sample containing the analyte molecule, with the supramolecular structure for a time period. In some embodiments, the time period is from about Is to about 24 hrs.

[0008] In some embodiments, for a nucleic acid described herein, the barcode of each binding molecule of the one or more binding molecules is a single stranded or double stranded nucleic acid having a unique sequence. In some embodiments, the at least one of the one or more reactive binders is cleavable, so as to separate the barcode from the affinity binder. In some embodiments, the tail section of each binding molecule of the one or more binding molecules further comprises a tail reactive group configured to link with the second reactive group and/or with the barcode from a different binding molecule of the one or more binding molecules. In some embodiments, linking the first binding molecule to the second reactive group comprises ligating a first tail reactive group of the first tail section with the second reactive group. In some embodiments, the process further comprises, prior to step (d), further comprising contacting the supramolecular structure with a reagent solution, thereby enabling separating the affinity binder and the portion of the first tail section from the first barcode. In some embodiments, the reagent solution comprises a low concentration of detergent, urea, formamide, or any combination thereof. In some embodiments, separating the portion of the first tail section from the first barcode is via strand displacement. In some embodiments, strand displacement is via a single- stranded DNA comprising a sequence complementary to the sequence of a portion of the tail section of the binding molecule.

[0009] In some embodiments, the process for forming a nucleic acid described herein further comprises binding the barcode with a protection binder wherein the protection binder is a single stranded nucleic acid of a sequence complementary to the sequence of the barcode. In some embodiments, the process further comprises providing a sample containing the analyte molecule, wherein the sample comprises a complex biological sample and the process provides for single-molecule sensitivity thereby increasing a dynamic range and quantitative capture of a range of molecular concentrations within the complex biological sample.

[00010] In some embodiments, the one or more analyte molecules comprises a protein, a peptide, a peptide fragment, a lipid, a DNA, a RNA, an organic molecule, an inorganic molecule, complexes thereof, or any combinations thereof. In some embodiments, each supramolecular structure is a nanostructure. In some embodiments, each core structure is a nanostructure. In some embodiments, the plurality of core molecules for each core structure are arranged into a pre-defmed shape and/or have a prescribed molecular weight. In some embodiments, the pre-defmed shape is configured to limit or prevent cross-reactivity with another supramolecular structure. In some embodiments, the plurality of core molecules for each core structure comprises one or more nucleic acid strands, one or more branched nucleic acids, one or more peptides, one or more small molecules, or combinations thereof. In some embodiments, each core structure independently comprises a scaffolded deoxyribonucleic acid (DNA) origami, a scaffolded ribonucleic acid (RNA) origami, a scaffolded hybrid DNA:RNA origami, a single-stranded DNA tile structure, a multi -stranded DNA tile structure, a single- stranded RNA origami, a multi -stranded RNA tile structure, hierarchically composed DNA or RNA origami with multiple scaffolds, a peptide structure, or combinations thereof. In some embodiments, the first reactive group comprises an amine, a thiol, a DBCO, a maleimide, a streptavidin, a biotin, an azide, an acrydite, aNHS-ester, a single stranded nucleic acid (RNA or DNA) of specific sequence, one or more polymers like PEG or polymerization initiators, or a combination thereof. In some embodiments, the second reactive group comprises a single stranded nucleic acid of a specific sequence. In some embodiments, the second reactive group comprises a terminal phosphate, amine, maleimide, or a combinations thereof such that it forms a chemical linkage with a single stranded nucleic acid. In some embodiments, the chemical linkage comprises a phosphodiester bond. In some embodiments, the first location and the second location are pre-determined or programmable on the core structure. [00011] Disclosed herein, in some embodiments, is a process for identifying an analyte molecule present in a sample, the process comprising: providing a supramolecular structure comprising: a core structure comprising a plurality of core molecules, a first reactive group linked to the core structure at a first location and configured to form a linkage with an analyte molecule, and a second reactive group linked to the core structure at a second location that is spaced apart from the first location by a prescribed distance, the second reactive group configured to form a linkage with one or more reporter molecules wherein each reporter molecule comprises i) an affinity binder configured to bind with the analyte molecule, ii) a tail section, and iii) at least one signaling molecules linked between the affinity binder and the tail section; contacting the supramolecular structure with the sample, such that the analyte molecule interacts with the first reactive group and is bound thereto; contacting a first solution comprising a first reporter molecule of the one or more reporter molecules with the supramolecular structure; detecting i) the supramolecular structure linked with the first reporter molecule via a first signaling molecule of the first reporter molecule, wherein a first affinity binder of the first reporter molecule is linked to the analyte molecule, and a first tail section of the first reporter molecule is linked to the second reactive group, or ii) the supramolecular structure not being linked to the first reporter molecule; when the supramolecular structure is linked with the first reporter molecule, separating the first reporter molecule from the supramolecular structure; contacting a second solution comprising a second reporter molecule of the one or more reporter molecules with the supramolecular structure, detecting i) the supramolecular structure linked with the second reporter molecule via a second signaling molecule of the second reporter molecule, wherein a second affinity binder of the second reporter molecule is linked to the analyte molecule, and a second tail section of the second reporter molecule is linked to the second reactive group, or ii) the supramolecular structure not being linked to the second reporter molecule; identifying the analyte molecule based on detecting the first and/or second reporter molecule being linked to the supramolecular structure. In some embodiments, the process, when the supramolecular structure is detected not being linked to the first reporter molecule, further comprises prior to step (e), tuning the second reporter molecule such that i) the second affinity binder is different from the first affinity binder, ii) the second tail section comprises a modified first tail section to increase an affinity strength with the second reactive molecule, or iii) a combination thereof. In some embodiments, the process comprises repeating steps (c)-(e) one or more times each with a different reporter molecule of the one or more reporter molecules, such that the identifying the analyte molecule is based on detecting the first reporter molecule, the second molecule, and/or any combination of one or more reporter molecules for the one or more times where steps (c)- (e) is repeated.

[00012] In some embodiments, the process for identifying an analyte molecule described herein further comprises quantifying the concentration of the analyte molecule in the sample. In some embodiments, the process further comprises identifying the detected analyte molecule. In some embodiments, the analyte molecule interacts with the first reactive group via a chemical bond. In some embodiments, the chemical bond comprises a covalent bond. In some embodiments, the chemical bond comprises a non-covalent bond. In some embodiments, the process comprises, prior to step (c), incubating the sample containing the analyte molecule, with the supramolecular structure for a time period. In some embodiments, the time period is from about Is to about 24 hrs. In some embodiments, the tail section of each reporter molecule of the one or more reporter molecules is a single stranded or double stranded nucleic acid. In some embodiments, tuning the first reporter molecule comprises increasing a length of a nucleic acid sequence of the tail section nucleic acid. In some embodiments, for a process described herein, the nucleic acid sequence of the tail section nucleic acid comprises from about 2 bases to about 10 bases. In some embodiments, tuning the first reporter molecule comprises changing a nucleic acid sequence of the tail section nucleic acid. In some embodiments, for a process described herein, the signaling molecule comprises a fluorophore, a redox molecule, or a magnetically active molecule. In some embodiments, the distance between the first location and the second location are spaced apart by a prescribed distance. In some embodiments, for a process described herein, the prescribed distance is about 1 nm to about 100 nm. In some embodiments, the process, prior to step (e), further comprises contacting the supramolecular structure with a reagent solution, thereby enabling separating the first reporting molecule from the supramolecular structure. In some embodiments, for a process described herein, the reagent solution comprises a mild detergent, urea, formamide, or any combination thereof. In some embodiments, for a process described herein, detecting the supramolecular structure being linked with the first reporter molecule is via a imaging the supramolecular structure. In some embodiments, the process further comprises the recording the detection of step (d) and (g).

[00013] In some embodiments, for a process described herein, the sample comprises a complex biological sample and the process provides for single-molecule sensitivity thereby increasing a dynamic range and quantitative capture of a range of molecular concentrations within the complex biological sample. In some embodiments, for a process described herein, the one or more analyte molecules comprises a protein, a peptide, a peptide fragment, a lipid, a DNA, a RNA, an organic molecule, an inorganic molecule, complexes thereof, or any combinations thereof. In some embodiments, each supramolecular structure is a nanostructure. In some embodiments, for a process described herein, each core structure is a nanostructure. In some embodiments, the plurality of core molecules for each core structure are arranged into a pre-defmed shape and/or have a prescribed molecular weight. In some embodiments, the pre defined shape is configured to limit or prevent cross-reactivity with another supramolecular structure. In some embodiments, the plurality of core molecules for each core structure comprises one or more nucleic acid strands, one or more branched nucleic acids, one or more peptides, one or more small molecules, or combinations thereof. In some embodiments, each core structure independently comprises a scaffolded deoxyribonucleic acid (DNA) origami, a scaffolded ribonucleic acid (RNA) origami, a scaffolded hybrid DNA:RNA origami, a single- stranded DNA tile structure, a multi -stranded DNA tile structure, a single-stranded RNA origami, a multi -stranded RNA tile structure, hierarchically composed DNA or RNA origami with multiple scaffolds, a peptide structure, or combinations thereof. In some embodiments, the first reactive group comprises an amine, a thiol, a DBCO, a maleimide, a streptavidin, a biotin, an azide, an acrydite, a NHS-ester, a single stranded nucleic acid (RNA or DNA) of specific sequence, one or more polymers like PEG or polymerization initiators, or a combination thereof. In some embodiments, the second reactive group comprises a single stranded nucleic acid of a specific sequence. In some embodiments, the supramolecular structure further comprises an identifier molecule linked thereto. In some embodiments, the identifier molecule comprises DNA (e.g., of a specific sequence), RNA (e.g., of a specific sequence), fluorophore, or a combination thereof.

BRIEF DESCRIPTION OF THE DRAWINGS [00014] Specific embodiments of the disclosed devices, delivery systems, or methods will now be described with reference to the drawings. Nothing in this detailed description is intended to imply that any particular component, feature, or step is essential to the invention. [00015] FIG. 1 provides an exemplary depiction of a supramolecular structure and the related subcomponents.

[00016] FIG. 2 provides an exemplary depiction of an assembled three-arm nucleic acid junction based supramolecular structure and related subcomponents.

[00017] FIG. 3 provides an exemplary depiction of the individual subcomponents of the three- arm nucleic acid junction based supramolecular structure from FIG. 2. [00018] FIG. 4 provides an exemplary depiction of an assembled DNA origami based supramolecular structure and related subcomponents.

[00019] FIG. 5 provides an exemplary depiction of the individual subcomponents of the DNA origami based supramolecular structure from FIG. 4.

[00020] FIGS. 6 A and B provide exemplary depictions of the structure of a binding molecule as described herein.

[00021] FIGS. 7A-C provides an exemplary depiction of a scheme of the identification of analyte molecules in a sample a using supramolecular structure. FIG. 7A provides a scheme of steps 1 to 3. FIG. 7B provides a scheme of steps 4 to 6. FIG 7C. illustrates detailed binding process for steps 3 and 4.

[00022] FIG. 8A provides an exemplary depiction of the structure of the reporter molecule comprising nucleic acid tail of sequence E.

[00023] FIG. 8B provides an exemplary depiction of the structure of the reporter molecule comprising nucleic acid tail of sequence F.

[00024] FIG. 9A provides an exemplary array of supramolecular structures described herein for identifying analyte molecules according to the scheme in FIG. 9B.

[00025] FIG. 9B provides another exemplary depiction of a scheme of the identification of analyte molecules in a sample using a supramolecular structure.

DETAILED DESCRIPTION

[00026] Disclosed herein are structures and methods for identifying one or more analyte molecules. In some embodiments, one or more supramolecular structures are used to identify the one or more analyte molecules. In some embodiments, identifying an analyte molecule, via a structure and/or method disclosed herein, comprises correlating a specific combination of affinity binders that are configured to form a linkage with an analyte molecule. In some embodiments, as described herein, the affinity binders are provided with binding molecules that comprise barcodes corresponding to the affinity binders. In some embodiments, the affinity binders are provided with reporter molecules comprising signaling molecules. In some embodiments, the one or more analyte molecules are provided in a sample. In some embodiments, the sample is contacted with the one or more supramolecular structures, thereby enabling one or more analyte molecules to bind with a corresponding supramolecular structure.

[00027] In one aspect for identifying an analyte molecule, a nucleic acid is formed on a supramolecular structure according to a method described herein, wherein the nucleic acid is formed with a sequence that corresponds to one or more analyte molecules. In some embodiments, the sequence is formed via successive linkage of a set of two or more segments to a corresponding supramolecular structure. In some embodiments, each segment comprises a nucleic acid barcode . In some embodiments, each nucleic acid barcode corresponds to an affinity binder. In some embodiments, the set of two or more segments identifies the analyte molecule. In some embodiments, a single segment identifies the analyte molecule. In some embodiments, the one or more barcodes are provided via one or more corresponding binding molecules. In some embodiments, each binding molecule comprises an affinity binder configured to bind with an analyte molecule, and a tail section configured to link with another portion of the supramolecular structure. In some embodiments, the tail section helps the corresponding affinity binder to bind with the analyte molecule where there is a weak affinity therebetween. In some embodiments the nucleic acid sequence is formed via successive iterations of allowing an interaction of a binding molecule with a supramolecular structure, followed by a removal of a portion of the binding molecule, leaving a corresponding barcode as part of the nucleic acid sequence. Accordingly, in some embodiments, an analyte molecule is identified based on the nucleic acid sequence formed on the supramolecular structure after one or more of such iterations.

[00028] In some embodiments, the segment of the barcode comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. In some embodiments, the segment of the barcode comprises between 1 and 10, between 2 and 9, between 3 and 8, between 4 and 7, between 5 and 6, or between 5 and 8. In another aspect for detecting an analyte molecule, one or more iterations of providing a reporter molecule to interact with an analyte molecule and supramolecular structure followed by removing the reporter molecule is performed, wherein each occurrence of an interaction between a reporter molecule and analyte molecule and supramolecular structure is recorded. In some embodiments, each reporter molecule comprises an affinity binder configured to bind with an analyte molecule, a signaling molecule to enable identification of an interaction with the supramolecular structure (and analyte molecule), and a tail section configured to link with a portion of the supramolecular structure. In some embodiments, the tail section and the affinity binder enable for co-operative binding to compensate for weak affinity between the affinity binder and an analyte molecule. In some embodiments, where no interaction (e.g., linkage) between a reporter molecule and supramolecular structure is identified, the reporter molecule may be tuned by changing the affinity binder, and/or modifying the tail section, to help boost an affinity of the reporter molecule to the analyte molecule and/or supramolecular structure. Accordingly, in some embodiments, an analyte molecule is identified based on the recorded occurrences of one or more reporter molecule binding with an analyte molecule, wherein the specific combination of different reporter molecules found to have bound with the analyte molecule enables for the detection and identification of said analyte molecule.

Sample

[00029] In some embodiments, the sample comprises an aqueous solution comprising protein, peptides, peptide fragments, lipids, DNA, RNA, organic molecules, inorganic molecules, complexes thereof, or any combinations thereof. In some embodiments, the analyte molecules in the sample comprise protein, peptides, peptide fragments, lipids, DNA, RNA, organic molecules, inorganic molecules, complexes thereof, or any combinations thereof. In some embodiments, the analyte molecules comprise intact proteins, denatured proteins, partially or fully degraded proteins, peptide fragments, denatured nucleic acids, degraded nucleic acid fragments, complexes thereof, or combinations thereof. In some embodiments, the sample is obtained from tissue, cells, the environment of tissues and/or cells, or combinations thereof. In some embodiments, the sample comprises tissue biopsy, blood, blood plasma, urine, saliva, a tear, cerebrospinal fluid, extracellular fluid, cultures cells, culture media, discarded tissue, plant matter, synthetic proteins, bacterial, viral samples, fungal tissue, or combinations thereof. In some embodiments, the sample is isolated from a primary source such as cells, tissue, bodily fluids (e.g., blood), environmental samples, or combinations thereof, with or without purification. In some embodiments, the cells are lysed using a mechanical process or other cell lysis methods (e.g., lysis buffer). In some embodiments, the sample is filtered using a mechanical process (e.g., centrifugation), micron filtration, chromatography columns, other filtration methods, or combinations thereof. In some embodiments, the sample is treated with one or more enzymes to remove one or more nucleic acids or one or more proteins. In some embodiments, the sample comprises intact proteins, denatured proteins, partially or fully degraded proteins, peptide fragments, denatured nucleic acids or degraded nucleic acid fragments. In some embodiments, the sample is collected from one or more individual persons, one or more animals, one or more plants, or combinations thereof. In some embodiments, the sample is collected from an individual person, animal and/or plant having a disease or disorder that comprises an infectious disease, an immune disorder, a cancer, a genetic disease, a degenerative disease, a lifestyle disease, an injury, a rare disease, an age-related disease, or combinations thereof.

Supramolecular Structure

[00030] In some embodiments, the supramolecular structure is a programmable structure that can spatially organize molecules. In some embodiments, the supramolecular structure comprises a plurality of molecules linked together. In some embodiments, the plurality of molecules of the supramolecular structure interact with at least some of each other. In some embodiments, the supramolecular structure comprises a specific shape. In some embodiments, the supramolecular nanostructure comprises a prescribed molecular weight based on the plurality of molecules of the supramolecular structure. In some embodiments, the supramolecular structure is a nanostructure. In some embodiments the plurality of molecules is linked together through a bond, a chemical bond, a physical attachment, or combinations thereof. In some embodiments, the supramolecular structure comprises a large molecular entity, of specific shape and molecular weight, formed from a well-defined number of smaller molecules interacting specifically with each other. In some embodiments, the structural, chemical, and physical properties of the supramolecular structure are explicitly designed. In some embodiments, the supramolecular structure comprises a plurality of subcomponents that are spaced apart according to a prescribed distance. In some embodiments, at least a portion of the supramolecular structure is rigid. In some embodiments, at least a portion of the supramolecular structure is semi-rigid. In some embodiments, at least a portion of the supramolecular structure is flexible.

[00031] FIG. 1 provides an exemplary embodiment of a supramolecular structure (30) comprising a core structure (9), a first reactive group (1), a second reactive group (2), and an anchor molecule (10). In some embodiments, the supramolecular structure comprises one or more first reactive groups (1), one or more second reactive groups (2), and/or optionally one or more anchor molecules (10). In some embodiments, the supramolecular structure does not comprise an anchor molecule. In some embodiments, the supramolecular structure is a polynucleotide structure.

[00032] In some embodiments, the core structure (9) comprises one or more core molecules linked together. In some embodiments, the one or more core molecules comprise 2, 3, 4, 5, 6,

7, 8, 9, 10, 20, 50, 100, 200 or 500 unique molecules that are linked together. In some embodiments, the one or more core molecules comprises from about 2 unique molecules to about 1000 unique molecules. In some embodiments, the one or more core molecules interact with each other and define the specific shape of the supramolecular structure. In some embodiments, the plurality of core molecules interacts with each other through reversible non- covalent interactions. In some embodiments, the specific shape of the core structure is a three- dimensional (3D) configuration. In some embodiments, the one or more core molecules provide a specific molecular weight. In some embodiments, the core structure 9 is a nanostructure. In some cases, the one or more core molecules comprise one or more nucleic acid strands (e.g., DNA, RNA, unnatural nucleic acids), one or more branched nucleic acids, one or more peptides, one or more small molecules, or combinations thereof. In some embodiments, the core structure 9 comprises a polynucleotide structure. In some embodiments, at least a portion of the core structure 9 is rigid. In some embodiments, at least a portion of the core structure 9 is semi-rigid. In some embodiments, at least a portion of the core structure 9 is flexible. In some embodiments, the core structure 9 comprises a scaffolded deoxyribonucleic acid (DNA) origami, a scaffolded ribonucleic acid (RNA) origami, a scaffolded hybrid DNA / RNA origami, a single-stranded DNA tile structure, a multi-stranded DNA tile structure, a single-stranded DNA origami, a single-stranded RNA origami, a single-stranded RNA tile structure, a multi -stranded RNA tile structures, a hierarchically composed DNA and/or RNA origami with multiple scaffolds, a peptide structure, or combinations thereof. In some embodiments, the DNA origami is scaffolded. In some embodiments, the RNA origami is scaffolded. In some embodiments, the hybrid DNA/RNA origami is scaffolded. In some embodiments, the core structure comprises a DNA origami, RNA origami, or hybrid DNA/RNA origami that comprises a prescribed two-dimensional (2D) or 3D shape.

[00033] As used herein, the term “nucleic acid origami” generally refers to a nucleic acid construct comprising an engineered tertiary (e.g., folding and relative orientation of secondary structures) or quaternary structure (e.g., hybridization between strands that are not covalently linked to each other) in addition to the naturally-occurring secondary structure (e.g., helical structure) of nucleic acid(s). A nucleic acid origami may include DNA, RNA, PNA, modified or non-natural nucleic acids, or combinations thereof. A nucleic acid origami can include a scaffold strand. The scaffold strand can be circular (i.e., lacking a 5’ end and 3’ end) or linear (i.e., having a 5’ end and/or a 3’ end). A nucleic acid origami may include a plurality of oligonucleotides that hybridize via sequence complementarity to produce the engineered structuring of the origami particle. For example, the oligonucleotides can hybridize to a scaffold strand and/or to other oligonucleotides. A nucleic acid origami may comprise sections of single-stranded or double-stranded nucleic acid, or combinations thereof. Exemplary nucleic acid origami structures may include nanotubes, nanowires, cages, tiles, nanospheres, blocks, and combinations thereof.

[00034] As shown in FIG 1, in some embodiments, the core structure 9 is configured to be linked to a first reactive group 1, a second reactive group 2, or a combination thereof. In some embodiments, the first reactive group 1 and/or the second reactive group 2 are immobilized with respect to the core nanostructure 9 when linked thereto. In some embodiments, any number of the one or more core molecules comprises one or more core linkers 6 and 8, configured to form a linkage with a first reactive group 1 and/or a second reactive group 2, respectively. In some embodiments, any number of the one or more core molecules are configured to be linked with one or more core linkers 6 and 8 that are configured to form a linkage with a first reactive group 1 and/or a second reactive group 2, respectively. In some embodiments, one or more core linkers are linked to one or more core molecules through a chemical bond. In some embodiments, at least one of the one or more core linkers comprises a DNA sequence domain.

[00035] In some embodiments, the core structure 9 is linked to 1) a first reactive group 1 at a prescribed first location on the core structure, 2) a second reactive group 2 at a prescribed second location on the core structure, 3) an anchor molecule 10 at a prescribed third location on the core structure, and/or 4) an identifier molecule at a prescribed fourth location on the core structure. In some embodiments, a specified first core linker 6 is disposed at the first location on the core structure, and a specified second core linker 8 is disposed at the second location on the core structure. In some embodiments, one or more core molecules at the first location are modified to form a linkage with the first core linker 6. In some embodiments, the first core linker 6 is an extension of the core structure 9. In some embodiments, one or more core molecules at the second location is modified to form a linkage with the second core linker 8. In some embodiments, the second core linker 8 is an extension of the core structure 9. In some embodiments, the 3D shape of the core structure 9 and relative distances of the first and second locations are specified to maximize the intramolecular interactions between the first reactive group 1 and the second reactive group 2. In some embodiments, the 3D shape of the core structure 9 and relative distances of the first and second locations are specified to obtain a desired distance between the first reactive group 1 and the second reactive group 2, so as to maximize the intramolecular interactions between the first reactive group 1 and the second reactive group 2.

[00036] As described herein, in some embodiments, the distance between the first reactive group 1 and the second reactive group 2 is about 1 nm, 2 nm, 3 nm, 4 nm, 5 nm, 6 nm, 10 nm, 12 nm, 15 nm, 20 nm, 30 nm, 40 nm, 50 nm, or 60 nm. In some embodiments, the distance between the first reactive group 1 and the second reactive group 2 is about 1 nm to about 60 nm. In some embodiments, the distance between the first reactive group 1 and the second reactive group 2 is about 1 nm to about 2 nm, about 1 nm to about 5 nm, about 1 nm to about 10 nm, about 1 nm to about 20 nm, about 1 nm to about 40 nm, about 1 nm to about 60 nm, about 2 nm to about 5 nm, about 2 nm to about 10 nm, about 2 nm to about 20 nm, about 2 nm to about 40 nm, about 2 nm to about 60 nm, about 3 nm to about 5 nm, about 3 nm to about 10 nm, about 3 nm to about 20 nm, about 3 nm to about 40 nm, about 3 nm to about 60 nm, about 4 nm to about 5 nm, about 4 nm to about 10 nm, about 4 nm to about 20 nm, about 4 nm to about 40 nm, about 4 nm to about 60 nm, about 5 nm to about 10 nm, about 5 nm to about 20 nm, about 5 nm to about 40 nm, about 5 nm to about 60 nm, about 6 nm to about 10 nm, about 6 nm to about 20 nm, about 6 nm to about 40 nm, about 6 nm to about 60 nm, about 7 nm to about 10 nm, about 7 nm to about 20 nm, about 7 nm to about 40 nm, about 7 nm to about 60 nm, about 8 nm to about 10 nm, about 8 nm to about 20 nm, about 8 nm to about 40 nm, about 8 nm to about 60 nm, about 9 nm to about 10 nm, about 9 nm to about 20 nm, about 9 nm to about 40 nm, about 9 nm to about 60 nm, about 10 nm to about 20 nm, about 10 nm to about 40 nm, about 10 nm to about 60 nm, about 20 nm to about 40 nm, about 20 nm to about 60 nm, or about 40 nm to about 60 nm, including increments therein. In some embodiments, the distance between the capture molecule 2 and detector molecule 1 is about 1 nm, about 2 nm, about 5 nm, about 10 nm, about 20 nm, about 40 nm, about 50 nm, or about 60 nm. In some embodiments, the distance between the first reactive group 1 and the second reactive group 2 is at least about 1 nm, about 2 nm, about 3 nm, about 4 nm, about 5 nm, about 6 nm, about 7 nm, about 8 nm, about 9 nm, about 10 nm, about 20 nm, about 40 nm, 50 nm, or 60 nm. In some embodiments, the distance between the first reactive group 1 and the second reactive group 2 is at most about 2 nm, about 5 nm, about 10 nm, about 20 nm, about 40 nm, 50 nm, or about 60 nm.

[00037] In some embodiments, the first, second, and fourth locations are disposed on a first side of the core structure 9, and the third location is disposed on a second side of the core structure 9.

[00038] In some embodiments, the first reactive group 1 comprises an antigen, a NHS ester, a maleimide, a biotin, an amine, a thiol, a DBCO, a maleimide, a streptavidin, an azide, an acrydite, a NHS-ester, a single stranded nucleic acid (RNA or DNA) of specific sequence, one or more polymers like PEG or polymerization initiators, or a combination thereof. In some embodiments, the first reactive group is configured to make a covalent bond with an arbitrary analyte molecule in a sample. In some embodiments, first reactive group 1 is configured to make a noncovalent bond with an arbitrary analyte molecule in a sample. In some embodiments, the first reactive group 1 is not randomly disposed on the core structure. In some embodiments, the location of the first reactive group 1 is pre-determined on the core structure. In some embodiments, the location of the first reactive group 1 is programmable at a molecular level on the core structure.

[00039] In some embodiments, the second reactive group 2 is linked to the core structure at the second location and configured to form a linkage with one or more reporter molecules. In some embodiments, the second reactive group 2 comprises a single stranded nucleic acid of a specific sequence (for example, B), thus a nucleic acid of a complementary sequence (for example, B’) can hybridize with. In some embodiments, the second reactive group 2 comprises a terminal phosphate, amine, maleimide, or a combinations thereof. In some embodiments, the second reactive group 2 forms a chemical linkage with a single stranded nucleic acid . In some embodiments, the second reactive group 2 forms a covalent linkage with a single stranded nucleic acid. In some embodiments, the second reactive group 2 and a single stranded nucleic acid are ligated. In some embodiments, the second reactive group 2 is not randomly disposed on the core structure. In some embodiments, the location of the second reactive group 2 is pre determined on the core structure. In some embodiments, the location of the second reactive group 2 is programmable at a molecular level on the core structure.

[00040] As used herein, the term “ligated” refers to the joining of two DNA fragments through the formation of a phosphodiester bond. An enzyme known as a DNA ligase catalyzes the formation of two covalent phosphodiester bonds between the 3’ hydroxyl group of one nucleotides and the 5’ phosphate group of another in an ATP dependent reaction.

[00041] In some embodiments, the identifier molecule 11 comprises a single stranded nucleic acid of a specific sequence and/or at least one fluorophore molecule. In some embodiments, a single pair of a first reactive group 1 and corresponding second reactive group 2 is linked to the core structure 9. In some embodiments, a plurality of pairs of a first reactive group 1 and corresponding second reactive group 2 are linked to a core structure 9. In some embodiments, the plurality of pairs of a first reactive group 1 and corresponding second reactive group 2 are spaced apart from each other to minimize cross-talk, i.e. minimizing the first reactive group 1 and/or the second reactive group 2 from a first pair interacting with the first reactive group 1 and/or the second reactive group 2 from a second pair.

[00042] In some embodiments, each component of the supramolecular structure may be independently modified or tuned. In some embodiments, modifying one or more of the components of the supramolecular structure may modify the 2D and 3D geometry of the supramolecular structure itself. In some embodiments, modifying one or more of the components of the supramolecular structure may modify the 2D and 3D geometry of the core structure. In some embodiments, such capability for independently modifying the components of the supramolecular nanostructure enables precise control over the organization of one or more supramolecular structures on solid surfaces (e.g., planar surfaces or microparticles) and 3D volumes (e.g., within a hydrogel matrix). [00043] As shown in FIG. 1, in some embodiments, the first reactive group 1 is linked to the core structure 9 through a first linker system. In some embodiments, the first linker system forms a linkage with the first reactive group 1, and the first linker system forms a linkage with the core structure 9. In some embodiments, the first linker system comprises a first linker 5 and a first bridge 3. In some embodiments, the first linker 5 comprises a reactive molecule. In some embodiments, the first linker 5 comprises a reactive molecule comprising an amine, a thiol, a DBCO, a NHS ester, a maleimide, an azide, an acrydite, a photocleavable linker, a single stranded nucleic acid (e.g., RNA or DNA) of specific sequence, a double stranded nucleic acid, or a polymer (e.g., polyethylene glycol (PEG) or one or more polymerization initiators). In some embodiments, the first linker 5 comprises a DNA sequence domain. In some embodiments, the second linker comprises a DNA sequence domain. In some embodiments, the first bridge 3 comprises a polymer. In some embodiments, the first bridge 3 comprises a polymer that comprises a nucleic acid (e.g., DNA or RNA) of a specific sequence. In some embodiments, the first bridge 3 comprises a polymer such as PEG. In some embodiments, the first linker 5 is attached to the first bridge 3 at a first terminal end thereof, and the first reactive group 1 is attached to the first bridge 3 at a second terminal end thereof. In some embodiments, the first linker 5 is attached to the first bridge 3 via a chemical bond. In some embodiments, the first linker 5 is attached to the first bridge 3 via a physical attachment.

[00044] In some embodiments, the first linker system is linked to the core structure 9 through a linkage between the first linker 5 and the first core linker 6. In some embodiments, as described herein, the first core linker 6 is disposed at a first location on the core structure 9. In some embodiments, the first linker 5 and first core linker 6 are linked together through a chemical bond. In some embodiments, the first linker 5 and first core linker 6 are linked together through a covalent bond. In some embodiments, the linkage between the first linker 5 and first core linker 6 is reversible upon being subjected to a trigger. In some embodiments the trigger signal comprises an optical signal. In some embodiments, the trigger signal comprises an electrical signal, microwave signal, ultraviolet illumination, visible illumination or near infra-red illumination.

[00045] As shown in FIG. 1, in some embodiments, the second reactive group 2 is linked to the core structure 9 through a second linker system. In some embodiments, the second linker system forms a linkage with the second reactive group 2, and the second linker system forms a linkage with the core structure 9. In some embodiments, the second linker system comprises a second linker 7 and a second bridge 4. In some embodiments, the second linker 7 comprises a reactive molecule. In some embodiments, the second linker 7 comprises a reactive molecule comprising an amine, a thiol, a DBCO, a NHS ester, a maleimide, biotin, an azide, an acrydite, a single stranded nucleic acid (e.g., RNA or DNA) of specific sequence, or a polymer (e.g., polyethylene glycol (PEG) or one or more polymerization initiators). In some embodiments, the second linker 7 comprises a DNA sequence domain. In some embodiments, the second bridge 4 comprises a polymer that comprises a nucleic acid (DNA or RNA) of a specific sequence. In some embodiments, the second bridge 4 comprises a polymer such as PEG. In some embodiments, the second linker 7 is attached to the second bridge 4 at a first terminal end thereof, and the second reactive group 2 is attached to the second bridge 4 at a second terminal end thereof. In some embodiments, the second linker 7 is attached to the second bridge 4 via a chemical bond. In some embodiments, the second linker 7 is attached to the second bridge 4 via a physical attachment.

[00046] In some embodiments, the second linker system is linked to the core structure 9 through a linkage between the second linker 7 and the second core linker 8. In some embodiments, as described herein, the second core linker 8 is disposed at a second location on the core structure 9. In some embodiments, the second linker 7 and second core linker 8 are linked together through a chemical bond. In some embodiments, the second linker 7 and second core linker 8 are linked together through a covalent bond. In some embodiments, the linkage between the second linker 7 and second core linker 8 is reversible upon being subjected to a trigger. In some embodiments the trigger signal comprises an optical signal. In some embodiments, the trigger signal comprises an electrical signal, microwave signal, ultraviolet illumination, visible illumination or near infra-red illumination.

Three arm nucleic acid junction based supramolecular structure

[00047] FIGS. 2 and 3 provide an exemplary depiction of a supramolecular structure 30 comprising a three arm nucleic acid junction and related subcomponents. FIG. 2 provides the complete supramolecular structure, while FIG. 3 provides the subcomponents that make up the supramolecular structure from FIG. 2. In some embodiments, the subcomponents of the supramolecular structure comprise total four DNA strands (ref. characters 20-23), including two reactor strands (20 & 23) and two core strands (21 & 22). The references characters 1-11 in FIGS. 2and 3 correspond to the respective components as provided with the same reference characters in FIG. 1.

[00048] As shown in FIGS. 2 and 3, in some embodiments of a supramolecular structure, the core structure comprises two strands, a first core strand 21 and a second core strand 22 that each comprise partially complementary DNA sequence domains labelled A and A, respectively. [00049] In some embodiments, the first core strand 21 of the core structure comprises a first core linker 6 comprising a DNA sequence domain. In some embodiments, the first core strand 21 comprises the DNA sequence domain labelled as “A” in FIGS. 2 and 3, which is separated from the first core linker 6 by an unstructured DNA region. In some embodiments, the unstructured DNA region comprises a polymer spacer. In some embodiments, the polymer spacer comprises a nucleic acid (DNA or RNA) of a specific sequence. In some embodiments, the polymer spacer comprises a polymer such as PEG.

[00050] In some embodiments, the first core linker 6 is complementary to a first linker 5 on the first reactor strand 20. In some embodiments, the first reactor strand 20 comprises a DNA strand comprising the first linker 5 and a first bridge 3 at either end of said first reactor strand 20. In some embodiments, the first linker 5 comprises a DNA sequence domain. In some embodiments, the first bridge 3 comprises a DNA sequence domain. In some embodiments, the first reactor strand 20 further comprises a unique first reactive group at the end of the first bridge 3. In some embodiments, the first reactive group 1 is covalently bound to the first bridge 3. In some embodiments, the first reactive group 1 comprises an antigen, a NHS ester, a maleimide, biotin, or combinations thereof.

[00051] In some embodiments, the second core strand 22 of the core structure comprises a second core linker 8 comprising a DNA sequence domain. In some embodiments, the second core strand 22 comprises the DNA sequence domain labelled as “A” in FIGS. 2 and 3, which is separated from the second core linker 8 by an unstructured DNA region. In some embodiments, the unstructured DNA region comprises a polymer spacer. In some embodiments, the polymer spacer comprises a nucleic acid (DNA or RNA) of a specific sequence. In some embodiments, the polymer spacer comprises a polymer such as PEG.

[00052] In some embodiments, the second core linker 8 is complementary to a second linker 7 on the second reactor strand 23. In some embodiments, the second reactor strand 23 comprises a DNA strand comprising the second linker 7 and a second bridge 4. In some embodiments, the second linker 7 comprises a DNA sequence domain. In some embodiments, the second bridge 4 comprises a DNA sequence domain. In some embodiments, the second reactor strand 23 further comprises a unique second reactive group 2 at the end of the second bridge 4.

[00053] In some embodiments, the second bridge 4 is complementary to a portion of a tail section of a binding molecule. In some embodiments, the portion of the tail section of a binding molecule is a DNA sequence domain. In some embodiments, the second reactive group 2 is bound to the second bridge 4. In some embodiments, the second reactive group 2 is covalently bound to the second bridge 4. [00054] In some embodiments, each of the different DNA domain sequences (reference character 3, 4, 5, 6, 7, 8, 9, A, A,) independently comprise nucleic acid sequences from about 2 nucleotides to about 80 nucleotides.

DNA origami based supramolecular structure

[00055] FIGS. 4 and 5 provide an exemplary depiction of a supramolecular structure 30 comprising a DNA origami and related subcomponents. FIG. 4 provides the complete supramolecular structure, while FIG. 5 provides the subcomponents that make up the supramolecular structure from FIG. 4. In some embodiments, the subcomponents of the supramolecular structure comprise a DNA origami 9 as a core structure and four (4) DNA strands (ref. characters 20-23). The references characters 1-11 in FIGS. 4 and 5 correspond to the respective components as provided with the same reference characters in FIG. 1.

[00056] In some embodiments, the core structure 9 comprises a scaffolded DNA origami, wherein a circular ssDNA molecule, called “scaffold” strand, is folded into a predefined 2D or 3D shape by interacting with 2 or more short ssDNA, called “staple” strands, which interact with specific sub-sections of the ssDNA “scaffold” strand.

[00057] As shown in FIGS. 4 and 5, in some embodiments of a supramolecular structure, the core structure 9 comprises a DNA origami. In some embodiments, the core structure 9 comprises a first core linker 6 comprising a DNA sequence domain. In some embodiments, the first core linker 6 is complementary to a first linker 5 on the first reactor strand 20. In some embodiments, the first reactor strand 20 comprises a DNA strand comprising the first reactive group 1 and the first linker 5 at either end of said first reactor strand 20. In some embodiments, the first linker 5 comprises a DNA sequence domain. In some embodiments, the first reactor strand 20 further comprises a nucleic acid (DNA or RNA) of a specific sequence in between the first reactive group 1 and the first linker 5. In some embodiments, the first reactor strand 20 comprises a polymer such as PEG in between the first reactive group 1 and the first linker 5. [00058] In some embodiments, the core structure 9 comprises a second core linker 8 comprising a DNA sequence domain. In some embodiments, the second core linker 8 is complementary to a second linker 7 on the second reactor strand 23. In some embodiments, the second reactor strand 23 comprises a DNA strand comprising the second linker 7 and a second reactive group 2 at either end of the second reactor section 23. In some embodiments, the second linker 7 comprises a DNA sequence domain. In some embodiments, the second reactor strand 23 further comprises a nucleic acid (DNA or RNA) of a specific sequence in between the second linker 7 and a second reactive group 2. In some embodiments, the specific sequence of the nucleic acid is complementary to a sequence of a portion of a tail section of a binding molecule.

Methods for Identifying Analyte Molecules

[00059] As described herein, in some embodiments, one or more supramolecular structures enable the identification of one or more analyte molecules in a sample. In some embodiments, as described herein, a plurality of supramolecular structures is provided in array to enable identification of a plurality of analyte molecules.

[00060] In some embodiments, methods described herein for identifying analytes in a sample provide a high-throughput and high-multiplexing capability by using a plurality of supramolecular structures. In some embodiments, the high-throughput and high-multiplexing capability provides high accuracy for analyte molecule identification and quantification. In some embodiments, methods described herein for identifying analytes in a sample are configured to characterize and/or identify biopolymers, including proteins molecules, quickly and at high sensitivity and reproducibility. In some embodiments, the plurality of supramolecular structures is configured to limit cross-reactivity associated errors.

[00061] In some embodiments, each core structure 9 of the plurality of supramolecular structures 30 is identical to one another. In some embodiments, the structural, chemical, and physical property of each supramolecular structure 30 is explicitly designed. In some embodiments, identical core structures 9 have a prescribed shape, size, molecular weight, prescribed number of a first reactive group 1 and a second reactive group 2, predetermined distance between a first reactive group 1 and a second reactive group 2 (as described herein), or combinations thereof, so as to limit the cross-reactivity between supramolecular structures 30. In some embodiments, the molecular weight of every core structure is identical and precise up to the purity of the core molecules 9. In some embodiments, each core structure 9 has at least one first reactive group 1 and at least one second reactive group 2.

[00062] In some embodiments, analyte molecules are provided in a sample (e.g., in a sample solution) that contacts a plurality of supramolecular structures 30, thereby enabling one or more analyte molecules to bind to a corresponding supramolecular structure 30. In some embodiments, the analyte molecules, as bound to the supramolecular structures 30, independently interact with one or more different affinity binders 604, wherein the interaction is driven primarily by intramolecular interaction. In some embodiments, the interaction between analyte molecules bound to the plurality of supramolecular structures 30 and different affinity binders 604 can be cooperatively induced by a programmable distance between a first reactive group 1 and a second reactive group 2 on the core structure 9 of the corresponding supramolecular structures 30. In some embodiments, as described herein, for each pair of a first reactive group 1 and a second reactive group 2 on a given supramolecular structure 30, the first reactive group 1 may specifically interact with one or more particular analyte molecules, which may in turn interact with one or more specific types of affinity binders 604, while the second reactive group 2 may specifically interact with a binding molecule or reporter molecule (as described herein), thereby leading to generating barcodes or light-sensitive response that can be correlated to the identification of an analyte molecule upon binding with the particular affinity binder 604. In some embodiments, a pair of a first reactive group 1 and a second reactive group 2 on a given supramolecular structure 30 is designed to interact with more than one analyte molecule in the sample and a corresponding affinity binder 604.

[00063] In some embodiments, each supramolecular structure is configured for single molecule sensitivity to ensure the identification of an analyte molecule in a typical complex biological sample. In some embodiments, single-molecule sensitivity comprises programmable location and structural features of a first reactive group 1 and a second reactive group 2 on a given supramolecular structure 30 at molecular level inducing a specific interaction between a particular analyte molecule and an affinity binder 604.

[00064] In some embodiments, the plurality of supramolecular structures 30 is provided in a solution. In some embodiments, the plurality of supramolecular structures 30 is attached to one or more substrates 714. In some embodiments, the plurality of supramolecular structures 30 is attached to one or more widgets. In some embodiments, the plurality of supramolecular structures 30 is attached to one or more solid substrates, one or more polymer matrices, one or more molecular condensates, or combinations thereof. In some embodiments, the one or more polymer matrices comprises one or more hydrogel particles. In some embodiments, the one or more polymer matrices comprises one or more hydrogel beads. In some embodiments, the one or more solid substrates comprises one or more planar substrates. In some embodiments, the one or more solid substrates comprises one or more microbeads. In some embodiments, the one or more solid substrates comprises one or more microparticles.

[00065] FIGS. 6A-7B depict an exemplary method and structure for identifying one or more analyte molecules. In some embodiments, one or more supramolecular structures 30 is provided (702). In some embodiments, the one or more supramolecular structures is provided on a substrate (714) as described herein. In some embodiments, the supramolecular structure 30 comprises a structure as described herein (e.g., see FIG. 1). For example, in some embodiments, the supramolecular structure (30) comprises a first reactive group (1), a second reactive group (2), and a core structure (9). In some embodiments, the supramolecular structures are each linked to the substrate via an anchor molecule (not shown) on the supramolecular structure (as described herein). In some embodiments, the supramolecular structures 30 are incubated with the substrate, thereby enabling the anchor molecules (or other attachment means) to link the supramolecular structures 30 to the substrate 714. In some embodiments, the incubation period is from about 30 seconds to about 24 hours. In some embodiments, the incubation period is from about 30 seconds to about 1 minute, from about 1 minute to about 5 minutes, from about 5 minutes to about 30 minutes, from about 30 minutes to about 1 hr, from about lhr to about 5 hours, from about 5 hours to about 12 hours, from about 12 hours to about 24 hours, from about 24 hours to about 48 hours. In some embodiments, the supramolecular structures 30 are arranged in an array, such that one or more locations on the array is identifiable with a corresponding supramolecular structure. In some embodiments, the supramolecular structures comprise an identifier molecule (not shown) to identify and/or label a supramolecular structure location. In some embodiments, the one or more supramolecular structures are placed in a solution.

[00066] In some embodiments, one or more analyte molecules are provided so as to enable interaction with the supramolecular structures. As described herein, the one or more analyte molecules may each comprise a protein. In some embodiments, the analyte molecules are provided in a sample. In some embodiments, the sample is provided as a solution. In some embodiments, the sample is contacted with the supramolecular structures (704), such that one or more supramolecular structures binds with a respective analyte molecule. In some embodiments, each supramolecular structure binds with a single analyte molecule. In some embodiments, a supramolecular structure may be configured to bind with two or more analyte molecules (not shown). In some embodiments, the supramolecular structure is incubated with the sample so that the analyte molecule interacts with the first reactive group (1) and forms a linkage with the first reactive group 1. In some embodiments, the incubation period is from about 30 seconds to about 24 hours. In some embodiments, the incubation period is from about 30 seconds to about 1 minute, from about 1 minute to about 5 minutes, from about 5 minutes to about 30 minutes, from about 30 minutes to about 1 hr, from about lhr to about 5 hours, from about 5 hours to about 12 hours, from about 12 hours to about 24 hours, from about 24 hours to about 48 hours. In some embodiments, the linkage between the first reactive group 1 and the analyte molecule comprises a chemical bond. In some embodiments, the linkage between the first reactive group 1 and the analyte comprises a covalent bond. In some embodiments, the linkage between the first reactive group 1 and the analyte molecule comprises an ionic bond. [00067] In some embodiments, after incubation of the sample with the supramolecular structures 30 and subsequent binding between one or more analyte molecules with a first reactive group 1 on a corresponding supramolecular structure 30, one or more binding molecules 602 are provided (706) for interaction with the analyte molecules and/or supramolecular structures 30. In some embodiments, the one or more binding molecules are provided in a solution that contacts the supramolecular structures 30. As described herein, in some embodiments, one or more iterations of providing a solution with binding molecules 602 to contact the supramolecular structure 30 is performed, wherein a partial denaturing step that separates at least a portion of binding molecules 602 linked with the supramolecular structures 30 and/or analyte molecules occurs in between each iteration of providing said solution with binding molecules 602. In some embodiments, the binding molecules provided with each iteration are different from each iteration (e.g., the first iteration comprises a first type of binding molecule for the one or more binding molecules, and a second iteration comprises a second type of binding molecule for the one or more binding molecules in the solution). In some embodiments, for each iteration, when two or more binding molecules are provided in the solution, said two or more binding molecules are of the same type (comprise same affinity binder, tail section). In some embodiments, for each iteration, when two or more binding molecules are provided in the solution, said two or more binding molecules are of different types.

[00068] In some embodiments of the present disclosure, each supramolecular structure 30 is a nanostructure. In some embodiments, each core structure 9 is a nanostructure. In some embodiments, the plurality of core molecules for each core structure 9 is arranged into a pre defined shape and/or have a prescribed molecular weight. In some embodiments, the pre defined shape is configured to limit or prevent cross-reactivity with another supramolecular structure 30. In some embodiments, the plurality of core molecules for each core structure 9 comprises one or more nucleic acid strands, one or more branched nucleic acids, one or more peptides, one or more small molecules, or combinations thereof. In yet other embodiments, each core structure 9 independently comprises a scaffolded deoxyribonucleic acid (DNA) origami, a scaffolded ribonucleic acid (RNA) origami, a scaffolded hybrid DNA:RNA origami, a single-stranded DNA tile structure, a multi -stranded DNA tile structure, a single-stranded RNA origami, a multi -stranded RNA tile structure, hierarchically composed DNA or RNA origami with multiple scaffolds, a peptide structure, or combinations thereof. In some embodiments, the supramolecular structure 30 comprises a DNA origami, a RNA origami, a DNA:RNA hybrid origami, or combinations thereof. [00069] In some embodiments, the first reactive group 1 is linked to the core structure 9 at the first location and configured to form a linkage with the analyte molecule. In some embodiments, the linkage between the first reactive group 1 and the core structure 9 is a chemical bond. In some embodiments, the linkage between the first reactive group 1 and the core structure 9 is a covalent bond. In some embodiments, the linkage between the first reactive group 1 and the core structure 9 is a noncovalent bond. In some embodiments, the linkage between the first reactive group 1 and the analyte molecule is a chemical bond. In some embodiments, the linkage between the first reactive group 1 and the analyte molecule is a covalent bond. In some embodiments, the linkage between the first reactive group 1 and the analyte molecule is a noncovalent bond.

[00070] In some embodiments, the first reactive group 1 comprises an antigen, a NHS ester, a maleimide, a biotin, an amine, a thiol, a DBCO, a maleimide, a streptavidin, an azide, an acrydite, a NHS-ester, a single stranded nucleic acid (RNA or DNA) of specific sequence, one or more polymers like PEG or polymerization initiators, or a combination thereof. In some embodiments, the first reactive group 1 makes a covalent bond with an arbitrary analyte molecule in a sample. In some embodiments, first reactive group 1 makes a noncovalent bond with an arbitrary analyte molecule in a sample. In some embodiments, the first reactive group 1 is not randomly disposed on the core structure 9. In some embodiments, the location of the first reactive group 1 is pre-determined on the core structure 9. In some embodiments, the location of the first reactive group 1 is programmable at a molecular level on the core structure 9.

[00071] In some embodiments, the second reactive group 2 is linked to the core structure 9 at the second location and configured to form a linkage with one or more binding molecules. In some embodiments, the second reactive group 2 comprises a terminal phosphate, amine, maleimide, or a combination thereof. In some embodiments, the second reactive group 2 forms a chemical linkage with a single stranded nucleic acid. In some embodiments, the second reactive group 2 forms a covalent linkage with a single stranded nucleic acid. In some embodiments, the second reactive group 2 and a single stranded nucleic acid are ligated. In some embodiments, the second reactive group 2 is not randomly disposed on the core structure 9. In some embodiments, the location of the second reactive group 2 is pre-determined on the core structure 9. In some embodiments, the location of the second reactive group 2 is programmable at a molecular level on the core structure 9.

[00072] FIGS. 6A-6B provide exemplary depictions of a binding molecule 602. With reference to FIG. 6A, in some embodiments, the binding molecule comprises an affinity binder (604) and a tail section (606). In some embodiments, the affinity binder (604) comprises an antibody, an antigen, a peptide, or an aptamer. In some embodiments, the tail section (606) comprises a first reactive binder (608), a second reactive binder (610), a barcode (612) that corresponds to the affinity binder (604), and/or a tail reactive group (614). In some embodiments, the first and/or second reactive binder each comprise a reactive cleavable binder. In some embodiments, the first and/or second reactive binder comprises a double stranded DNA, a thiol, a dithiol, a photocleavable linker, etc. In some embodiments, the first reactive binder (608) comprises a toehold (as known in the art). In some embodiments, the barcode (612) comprises a single stranded or double stranded nucleic acid. In some embodiments, the barcode (612) comprises a unique sequence corresponding to the affinity binder (604). In some embodiments, the tail reactive group (614) is configured to interact and link with the second reactive group of a supramolecular structure described herein. In some embodiments, the tail reactive group (614) comprises a single stranded nucleic acid (RNA or DNA) of specific sequence.

[00073] FIG. 6B provides a more detailed depiction of an exemplary binding molecule 602 described herein. In some embodiments, the binding molecule comprises i) an affinity binder 604 configured to bind with the analyte molecule and ii) a tail section 606 comprising a barcode 612 corresponding to the affinity binder 604. In some embodiments, as shown in FIG. 6B, the tail section 606 of the binding molecule 602 comprises a double stranded nucleic acid comprising a strand D1 and a strand D2. In some embodiments, the strand D1 comprises three continuously connected fragments of a single stranded DNA. In some embodiments, the three continuously connected fragment independently comprise a unique sequence of single stranded DNA. In some embodiments, the three continuously connected fragments of a single stranded DNA comprise sequences of B’, BC’ and B’ in order. In some embodiments, the strand D2 comprises two continuously connected fragments of a single stranded DNA. In some embodiments, the two continuously connected fragment of the strand D2 independently comprise a unique sequence of single stranded DNA. In some embodiments, the two continuously connected fragments of a single stranded DNA comprise sequences of B and BC in order. In some embodiment, the sequence B and BC are complementary to B’ and BC’, respectively. In some embodiments, as the strand D1 and the strand D2 bind, the fragment of sequence B’ of the three fragments of D1 is left as a single strand. In some embodiments, the sequence of BC of the strand D2 is the barcode 612 corresponding to the affinity binder 604. [00074] In some embodiments, as illustrated in FIG. 6B, the affinity binder 604 is linked to a short domain called the toeholds (“TH”). In some embodiments, TH comprises a unique sequence of nucleic acid. In some embodiments, the affinity binder 604 is linked to an end of TH and the other end of TH is linked to the sequence B’, of which double stranded fragment binding with D2, of the strand Dl.

[00075] In some embodiments, the affinity binder 604 has a high-affinity target binding. In some embodiments, the affinity binder has a weak-affinity target binding. In some embodiments, the affinity binder comprises a protein, a peptide, a peptide fragment, a lipid, a DNA, a RNA, an organic molecule, an inorganic molecule, complexes thereof, or any combinations thereof. In some embodiments, the affinity binder comprises proteins. In some embodiments, the protein comprises an antibody, an antigen, a peptide, or an aptamer. In some embodiments, a dissociation constant (K_d) value of the affinity binder is less than about 1 pM (picomolar). In some embodiments, the dissociation constant (K_d) value of the affinity binder is less than about 5 pM, about 4 pM, about 3 pM, about 2 pM, about 1 pM, about 0.9 pM, about 0.8 pM, about 0.7 pM, about 0.6 pM, about 0.5 pM, about 0.4 pM, about 0.3 pM, 0.2 pM, about 0.1 pM, about 0.05 pM, or about 0.01 pM. In some embodiments, the dissociation constant (K_d) value of the affinity binder is between about 0.01 pM and about 5 pM, between about 0.05 pM and about 4 pM, between about 0. 1 pM and about 3 pM, between about 0.2 pM and about 2 pM, between about 0.3 pM and about 1 pM, between about 0.4 pM and about 0.9 pM, between about 0.5 pM and about 0.8 pM, or between about 0.6 pM and about 0.7 pM.

[00076] With reference to FIG. 7A, reference character 706 provides a first iteration of contacting a first solution with one or more binding molecules (602) with the supramolecular structures. In some embodiments, at least two different binding molecules are provided with each iteration. In some embodiments, the supramolecular structures are incubated with the first solution containing the one or more binding molecules. Accordingly, in some embodiments, one or more analyte molecules bound to corresponding supramolecular structures forms a linkage to a respective affinity binder of a binding molecule. In some embodiments, prior to, after, simultaneously, or substantially simultaneously with an analyte molecule binding with a respective affinity binder, a tail section of the binding molecule forms a linkage with a second reactive group of a supramolecular structure (.e.g., via a tail reactive group). For example, as shown in FIG. 7A, a first binding molecule is depicted as forming a linkage (716) with the second reactive group of a supramolecular structure, via the respective tail section, and also forming a linkage between the analyte molecule and corresponding affinity binder (718). In some embodiments, the linkage between the second reactive group and the first binding molecule is hybridization between the tail reactive group and the second reactive group. For example, as described herein, in some embodiments, the second reactive group comprises a single stranded nucleic acid comprising a repeating sequence of B that is complementary to a sequence B’ (from a tail section of a binding molecule).

[00077] In further embodiments, the first barcode 612 of the strand D2 forms a chemical bond with the second reactive group 2. In some embodiments, the first barcode 612 of the strand D2 forms a covalent bond with the second reactive group 2. In some embodiments, the first barcode 612 of the strand D2 is ligated with the second reactive group 2.

[00078] In some embodiments, the next step (708) is separating the affinity binders 604 from the respective analyte molecules and separating a portion of the corresponding tail section 606 of a binding molecule from the barcode 612 of the same tail section. Accordingly, in some embodiments the barcode 612 remains linked to the second reactive group (720) while the remaining portion of the binding molecule and affinity binder are separated. In some embodiments, the affinity binder and corresponding portions of the binding molecules are separated from the respective analyte molecules and barcode sections, respectively, via a mild denaturing treatment. In some embodiments, the mild denaturing treatment comprises contacting the supramolecular structures with a low concentration of urea, detergent, formamide, or a combination thereof. In some embodiments, the strand D1 is separated from the strand D2 via binding with an invading DNA strand 722. In some embodiments, the invading strand 722 is a single stranded DNA of a complementary sequence to the sequence of D1 including TH region. In yet other embodiments, the strand D1 is separated from the strand D2 via strand displacement using the invading DNA strand 722. In further embodiments, the first barcode remains linked to the second reactive group as an elongated sequence. In additional embodiments, the elongated sequence comprises the sequence of B and BC. In some embodiments, a protecting strand of sequence BC’ hybridized with the first barcode of sequence BC.

[00079] As used herein, the term “hybridized” or “hybridization” refers to a phenomenon in which single-stranded DNA or RNA molecules bind to complementary strand DNA or RNA.

[00080] As used herein, the term “strand displacement” refers to a molecular tool to exchange one strand of DNA or RNA (output) with another strand (input). It is based on the hybridization of two complementary strands of DNA or RNA. It starts with a double-stranded DNA complex composed of the original strand and the protector strand. The original strand has an overhanging region the so-called “toehold” which is complementary to a third strand of DNA referred to as the “invading strand”. Accordingly, for example, the invading strand (722) is a sequence of single-stranded DNA (ssDNA) which is complementary to the original strand. The toehold regions initiate the process by allowing the complementary invading strand to hybridize with the original strand, creating a DNA complex composed of three strands of DNA. After the binding of the invading strand and the original strand occurred, branch migration of the invading domain then allows the displacement of the initial hybridized strand.

[00081] As described herein, upon separating affinity binders and portions of the binding molecules, in some embodiments, the supramolecular structures are contacted with another solution containing binding molecules (710) to repeat the steps of (706) and (708) again. In some embodiments, a protection strand 714 is further provided to hybridize with the single stranded barcode from the previous binding molecule that remains linked to the second reactive group for protection of the single stranded barcode strand from any unnecessary binding.

[00082] As depicted in (712) of FIG. 7B, after performing steps (706) to (710) a number of times, a nucleic acid having a specific sequence (724) will be formed on one or more of the supramolecular structures. As depicted, in some embodiments, the specific sequence will comprise the barcode from the binding molecules that interacted with the corresponding analyte molecule, wherein the tail sections of the binding molecules in subsequent iterations to the first iteration interact with the barcode added to the nucleic acid sequence from the respective previous iteration (e.g. second iteration encompasses tail section from second binding molecule forming a linkage with the barcode from the first binding molecule). In some embodiments, each nucleic acid sequence corresponds to a specific analyte molecule, thereby enabling its identification. For example, the supramolecular structures depicted in (712) of FIG. 7B all comprise a different sequence, signifying in some instances that the respective analyte molecules are configured to bind to a specific number of the affinity binders of the respective binding molecules. In some embodiments, the analyte molecules are identified via a DNA sequencing of the nucleic acid sequence. In some embodiments, the elongated barcode sequence is determined by nucleic acid sequencing. In some embodiments, the analyte molecule is identified upon determination of the elongated barcode sequence.

[00083] As described herein, in some embodiments, the supramolecular structures are provided in an array configuration. Accordingly, each nucleic acid sequence formed is analyzed to correlate the analyte molecule disposed at the given location of supramolecular structure array.

[00084] FIGS. 9A-9B provide another exemplary method and structure for identifying one or more analyte molecules. In some embodiments, one or more supramolecular structures 30 is provided (902). In some embodiments, the one or more supramolecular structures is provided on a substrate (914) as described herein. In some embodiments, the supramolecular structure 30 comprises a structure as described herein (e.g., see FIG. 1). For example, in some embodiments, the supramolecular structure (30) comprises a first reactive group (1), a second reactive group (2), and a core structure (9). In some embodiments, the supramolecular structures are linked to the substrate via an anchor molecule (not shown) on the supramolecular structure (as described herein). In some embodiments, the supramolecular structures 30 are incubated with the substrate, thereby enabling the anchor molecules (or other attachment means) to link the supramolecular structures 30 to the substrate 914. In some embodiments, the incubation period is from about 30 seconds to about 24 hours. In some embodiments, the incubation period is from about 30 seconds to about 1 minute, from about 1 minute to about 5 minutes, from about 5 minutes to about 30 minutes, from about 30 minutes to about 1 hr, from about lhr to about 5 hours, from about 5 hours to about 12 hours, from about 12 hours to about 24 hours, from about 24 hours to about 48 hours. In some embodiments, the supramolecular structures 30 are arranged in an array, such that one or more locations on the array is identifiable with a corresponding supramolecular structure. In some embodiments, the supramolecular structures comprise an identifier molecule (not shown) to identify and/or label a supramolecular structure location. In some embodiments, the one or more supramolecular structures are placed in a solution.

[00085] In some embodiments, one or more analyte molecules are provided so as to enable interaction with the supramolecular structures. As described herein, the one or more analyte molecules may each comprise a protein. In some embodiments, the analyte molecules are provided in a sample. In some embodiments, the sample is provided as a solution. In some embodiments, the sample is contacted with the supramolecular structures (904), such that one or more supramolecular structures each binds with an analyte molecule. In some embodiments, each supramolecular structure binds with a single analyte molecule. In some embodiments, a supramolecular structure may be configured to bind with two or more analyte molecules (not shown). In some embodiments, the supramolecular structure is incubated with the sample so that the analyte molecule interacts with the first reactive group (1) and forms a linkage with the first reactive group 1. In some embodiments, the incubation period is from about 30 seconds to about 24 hours. In some embodiments, the incubation period is from about 30 seconds to about 1 minute, from about 1 minute to about 5 minutes, from about 5 minutes to about 30 minutes, from about 30 minutes to about 1 hr, from about lhr to about 5 hours, from about 5 hours to about 12 hours, from about 12 hours to about 24 hours, from about 24 hours to about 48 hours. In some embodiments, the linkage between the first reactive group 1 and the analyte molecule comprises a chemical bond. In some embodiments, the linkage between the first reactive group 1 and the analyte comprises a covalent bond. In some embodiments, the linkage between the first reactive group 1 and the analyte molecule comprises an ionic bond.

[00086] In some embodiments, after incubation of the sample with the supramolecular structures 30 and subsequent binding between one or more analyte molecules with a first reactive group 1 on a corresponding supramolecular structure 30, one or more reporter molecules 802 are provided (906) for interaction with the analyte molecules and/or supramolecular structures 30. In some embodiments, the one or more reporter molecules are provided in a solution that contacts the supramolecular structures 30.

[00087] In some embodiments of the present disclosure, each supramolecular structure 30 is a nanostructure. In some embodiments, each core structure 9 is a nanostructure. In some embodiments, the plurality of core molecules for each core structure 9 is arranged into a pre defined shape and/or have a prescribed molecular weight. In some embodiments, the pre defined shape is configured to limit or prevent cross-reactivity with another supramolecular structure 30. In some embodiments, the plurality of core molecules for each core structure 9 comprises one or more nucleic acid strands, one or more branched nucleic acids, one or more peptides, one or more small molecules, or combinations thereof. In yet other embodiments, each core structure 9 independently comprises a scaffolded deoxyribonucleic acid (DNA) origami, a scaffolded ribonucleic acid (RNA) origami, a scaffolded hybrid DNA:RNA origami, a single-stranded DNA tile structure, a multi -stranded DNA tile structure, a single-stranded RNA origami, a multi -stranded RNA tile structure, hierarchically composed DNA or RNA origami with multiple scaffolds, a peptide structure, or combinations thereof. In some embodiments, the supramolecular structure 30 comprises a DNA origami, a RNA origami, a DNA:RNA hybrid origami, or combinations thereof.

[00088] In some embodiments, the first reactive group 1 is linked to the core structure 9 at the first location and configured to form a linkage with the analyte molecule. In some embodiments, the linkage between the first reactive group 1 and the core structure 9 is a chemical bond. In some embodiments, the linkage between the first reactive group 1 and the core structure 9 is a covalent bond. In some embodiments, the linkage between the first reactive group 1 and the core structure 9 is a noncovalent bond. In some embodiments, the linkage between the first reactive group 1 and the analyte molecule is a chemical bond. In some embodiments, the linkage between the first reactive group 1 and the analyte molecule is a covalent bond. In some embodiments, the linkage between the first reactive group 1 and the analyte molecule is a noncovalent bond.

[00089] In some embodiments, the first reactive group 1 comprises an antigen, a NHS ester, a maleimide, a biotin, an amine, a thiol, a DBCO, a maleimide, a streptavidin, an azide, an acrydite, a NHS-ester, a single stranded nucleic acid (RNA or DNA) of specific sequence, one or more polymers like PEG or polymerization initiators, or a combination thereof. In some embodiments, the first reactive group 1 makes a covalent bond with an arbitrary analyte molecule in a sample. In some embodiments, first reactive group 1 makes a noncovalent bond with an arbitrary analyte molecule in a sample. In some embodiments, the first reactive group 1 is not randomly disposed on the core structure 9 In some embodiments, the location of the first reactive group 1 is pre-determined on the core structure 9 In some embodiments, the location of the first reactive group 1 is programmable at a molecular level on the core structure 9

[00090] In some embodiments, the second reactive group 2 is linked to the core structure at the second location and configured to form a linkage with one or more reporter molecules. In some embodiments, the second reactive group 2 comprises a single stranded nucleic acid of a specific sequence (for example E’), thus a nucleic acid of a complementary sequence (for example E) can hybridize with. In some embodiments, the second reactive group 2 is not randomly disposed on the core structure. In some embodiments, the location of the second reactive group 2 is pre-determined on the core structure. In some embodiments, the location of the second reactive group 2 is programmable at a molecular level on the core structure.

[00091] In further embodiments, the distance D between the first location (of the supramolecular structure, where the first reactive group is linked to) and the second location (of the supramolecular structure, where the second reactive group is linked to) is pre-determined on the core structure. In some embodiments, the distance D between the first location and the second location is programmable on the core structure. In some embodiments, the distance D between the first location and the second location is short enough to cooperatively induce the interaction of the analyte molecule and the reporter molecule the distance D between the first reactive group 1 and the second reactive group 2 is about 1 nm, 2 nm, 3 nm, 4 nm, 5 nm, 6 nm, 10 nm, 12 nm, 15 nm, 20 nm, 30 nm, 40 nm, 50 nm, or 60 nm. In some embodiments, the distance D between the first reactive group 1 and the second reactive group 2 is about 1 nm to about 60 nm. In some embodiments, the distance D between the first reactive group 1 and the second reactive group 2 is about 1 nm to about 2 nm, about 1 nm to about 5 nm, about 1 nm to about 10 nm, about 1 nm to about 20 nm, about 1 nm to about 40 nm, about 1 nm to about 60 nm, about 2 nm to about 5 nm, about 2 nm to about 10 nm, about 2 nm to about 20 nm, about 2 nm to about 40 nm, about 2 nm to about 60 nm, about 3 nm to about 5 nm, about 3 nm to about 10 nm, about 3 nm to about 20 nm, about 3 nm to about 40 nm, about 3 nm to about 60 nm, about 4 nm to about 5 nm, about 4 nm to about 10 nm, about 4 nm to about 20 nm, about 4 nm to about 40 nm, about 4 nm to about 60 nm, about 5 nm to about 10 nm, about 5 nm to about 20 nm, about 5 nm to about 40 nm, about 5 nm to about 60 nm, about 6 nm to about 10 nm, about 6 nm to about 20 nm, about 6 nm to about 40 nm, about 6 nm to about 60 nm, about 7 nm to about 10 nm, about 7 nm to about 20 nm, about 7 nm to about 40 nm, about 7 nm to about 60 nm, about 8 nm to about 10 nm, about 8 nm to about 20 nm, about 8 nm to about 40 nm, about 8 nm to about 60 nm, about 9 nm to about 10 nm, about 9 nm to about 20 nm, about 9 nm to about 40 nm, about 9 nm to about 60 nm, about 10 nm to about 20 nm, about 10 nm to about 40 nm, about 10 nm to about 60 nm, about 20 nm to about 40 nm, about 20 nm to about 60 nm, or about 40 nm to about 60 nm, including increments therein. In some embodiments, the distance D between the capture molecule 2 and detector molecule 1 is about 1 nm, about 2 nm, about 5 nm, about 10 nm, about 20 nm, about 40 nm, about 50 nm, or about 60 nm. In some embodiments, the distance D between the first reactive group 1 and the second reactive group 2 is at least about 1 nm, about 2 nm, about 3 nm, about 4 nm, about 5 nm, about 6 nm, about 7 nm, about 8 nm, about 9 nm, about 10 nm, about 20 nm, about 40 nm, 50 nm, or 60 nm. In some embodiments, the distance D between the first reactive group 1 and the second reactive group 2 is at most about 2 nm, about 5 nm, about 10 nm, about 20 nm, about 40 nm, 50 nm, or about 60 nm.

[00092] In some embodiments, as shown in FIG. 8A, the reporter molecule 802 comprises i) an affinity binder 804 configured to bind with the analyte molecule, ii) a tail section 808, and iii) at least one signaling molecule 806 linked in between the affinity binder 804 and the tail section 808. In some embodiment, the tail section 808 of the reporter molecule comprises a single stranded nucleic acid of a specific sequence (for example, E) which is complementary to the sequence (for example, E’) of the second reactive group 2. In yet other embodiments, the at least one signaling molecule 806 comprises fluorescent moieties. In some embodiments, the at least one signaling molecule 806 comprises redox molecules. In some embodiments, the at least one signaling molecule 806 comprises magnetically active molecules. In some embodiments, the fluorescent moieties comprise fluorescent dye molecules, quantum dots, fluorescent carbon nanostructures, or combination thereof. Examples of fluorescent dye molecules include, but are not limited to, Fluorescein, Rhodamine, Oregon green, Texas red, cyanine, indocarbocyanine, oxacarbocyanine, thiacarbocyanine, merocyanine, pyridyloxazole, nitrobenzoxadiazole, benzoxadiazole, DRAQ5, DRAQ7, CyTRAK Orange, Alexa Fluor™

555, Alexa Fluor™ 647, BODIPY™ FL, Alexa Fluor™ 488, Alexa Fluor™ 546, Alexa Fluor™ 568, Alexa Fluor™ 594, or combination thereof. In some embodiments, the redox molecule comprises reactive oxygen species (ROS) such as superoxide, hydrogen peroxide, hydroxyl radical, nitric oxide, and peroxyni trite.

[00093] In some embodiments, the affinity binder has a high-affinity target binding. In some embodiments, the affinity binder has a weak-affmity target binding. In some embodiments, the affinity binder comprises a protein, a peptide, a peptide fragment, a lipid, a DNA, a RNA, an organic molecule, an inorganic molecule, complexes thereof, or any combinations thereof. In some embodiments, the affinity binder comprises proteins. In some embodiments, the protein comprises an antibody, an antigen, a peptide, or an aptamer. In some embodiments, a dissociation constant (K_d) value of the affinity binder is less than about 1 pM (picomolar). In some embodiments, the dissociation constant (K_d) value of the affinity binder is less than about 5 pM, about 4 pM, about 3 pM, about 2 pM, about 1 pM, about 0.9 pM, about 0.8 pM, about 0.7 pM, about 0.6 pM, about 0.5 pM, about 0.4 pM, about 0.3 pM, 0.2 pM, about 0.1 pM, about 0.05 pM, or about 0.01 pM. In some embodiments, the dissociation constant (K_d) value of the affinity binder is between about 0.01 pM and about 5 pM, between about 0.05 pM and about 4 pM, between about 0. 1 pM and about 3 pM, between about 0.2 pM and about 2 pM, between about 0.3 pM and about 1 pM, between about 0.4 pM and about 0.9 pM, between about 0.5 pM and about 0.8 pM, or between about 0.6 pM and about 0.7 pM.

[00094] With reference to FIG. 9B, reference character 906 provides a first iteration of contacting a first solution with one or more reporter molecules (802) with the supramolecular structures. In some embodiments, the one or more reporter molecules are the same type of reporter molecule. In some embodiments, the reporter molecules provided with each iteration are different from each iteration (e.g., the first iteration comprises a first type of reporter molecule for the one or more reporter molecules, and a second iteration comprises a second type of reporter molecule for the one or more reporter molecules in the solution). In some embodiments, for each iteration, when two or more reporter molecules are provided in the solution, said two or more reporter molecules are of the same type (comprise same affinity binder, tail section, and signaling molecule). In some embodiments, for each iteration, when two or more reporter molecules are provided in the solution, said two or more reporter molecules differ only via different signaling molecules thereon (e.g., different types of colors when imaging the supramolecular structure, as discussed herein). In some embodiments, for each iteration, when two or more reporter molecules are provided in the solution, said two or more reporter molecules are of different types.

[00095] In some embodiments, the supramolecular structures are incubated with the first solution containing the one or more reporter molecules so that the reporter molecule 802 interacts with the first reactive group 1 and the second reactive group 2. In some embodiments, the second reactive group 2 comprises a single stranded nucleic acid of a sequence (for example, E’) that is complementary to a sequence (for example, E). Accordingly, in some embodiments, the first reporter molecule 802 (from a first iteration of a solution with reporter molecules contacting the supramolecular structures) forms a linkage with the second reactive group 2 (sequence E’, for example) via a first tail section 808 (sequence E, for example) of the first reporter molecule 802. In some embodiments, the first reporter molecule 802 hybridizes with the second reactive group 2 (sequence E’) via a first tail section 808 (sequence E) of the first reporter molecule 802.

[00096] With reference to FIG. 9B, reference character 908 illustrates that the binding of the first reporter molecule 802 and the second reactive group 2 places the first reporter molecule 802 close to the first reactive group 1 on the core structure 9. In some embodiments, the location of the first reporter molecule 802 on the core structure 9 is determined upon binding to the second reactive group 2 on the core structure 9. In some embodiments, the distance D of the first reactive group 1 and the second reactive group 2 is pre-determined. In some embodiments, the distance D of the first reactive group 1 and the second reactive group 2 is programmable. In some embodiments, the second reactive group 2 is disposed close to the first reactive group 1 on the core structure. In yet other embodiments, the distance D between the first reactive group 1 and the second reactive group 2 is close enough to cooperatively induce the interaction of the first affinity binder 804 of the first reporter molecule 802 and an analyte molecule on a supramolecular structure, after the tail section 808 to form a linkage to the second reactive group 2 of the supramolecular structure. In still other embodiments, the distance D between the first reactive group 1 and the second reactive group 2 is pre-determined/programmed to place the first affinity binder 804 of the first reporter molecule 802 bound to the second reactive group 2 adjacent to the analyte molecule bound to the first reactive group 1, thus the interaction between the first affinity binder 804 and the analyte molecule is cooperatively induced and a linkage between them is formed. In still other embodiments, the interaction of the first affinity binder 804 of the first reporter molecule 802 and the analyte molecule bound to the first reactive group 1 is cooperatively induced by the linkage between the first reporter molecule 802 and the second reactive group 2, wherein the first affinity binder 804 is placed adjacent to the analyte molecule due to the programmable distance D between the first reactive group 1 and the second reactive group 2. In some embodiments, prior to, after, simultaneously, or substantially simultaneously with an analyte molecule binding with a respective affinity binder, a tail section of the reporter molecule forms a linkage with a second reactive group of a supramolecular structure (.e.g., via a tail reactive group).

[00097] In some embodiments, the solution containing reporter molecules (e.g., in a first iteration) is allowed to incubate with the supramolecular structures. In some embodiments, supramolecular structures are analyzed to identify (910) which supramolecular structures formed a linkage with a reporter molecule. In some embodiments, such identification is via imaging of the supramolecular structures. For example, in some embodiments, at least one first signaling molecule 806 is identified on the supramolecular structure upon binding the first reporter molecule 802 to the second reactive group 2 and the analyte molecule using one or more light-sensing imaging devices. Examples of light-sensing imaging device include, but are not limited to, a fluorescence microscope, a confocal microscope, Total Internal Reflection Microscopy (TIRF), or combination thereof. In some embodiments, as described herein, the supramolecular structures are provided on a substrate, such that the location of each supramolecular structure identified with a reporter molecule linked thereto is recorded (via a signal provided by the signaling molecules). In some embodiments, recording the detected reporter molecule linkage is via manual input, automated via a computing device, or a combination thereof. In some embodiments, as described herein, the supramolecular structure is provided in an array configuration, wherein each supramolecular structure is identified with an x, y coordinate system (see for example, 914).

[00098] In some embodiments, those supramolecular structures not identified with a binding molecule linked thereto are recorded as such (according to the location of the supramolecular structure on the substrate and/or array). For example, in some embodiments, as shown in FIG. 8B, the tail section 807 of the reporter molecule 803 comprises a single stranded nucleic acid of a specific sequence (for example, F) which is not complementary to the sequence (for example, E’) of the second reactive group 2. In some embodiments, the second reactive group 2 comprises a single stranded nucleic acid comprising a sequence of E’ that is not complementary to a sequence F. In some embodiments, the reporter molecule 803 does not form a linkage with the second reactive group 2 (sequence E’) via a tail section 807 (sequence F) of the reporter molecule 803. In yet other embodiments, the affinity binder 804 of the reporter molecule 803 interacts with the analyte molecule bound to the first reactive group and make a linkage between them. In yet other embodiments, the affinity binder 804 of the reporter molecule 803 interacts with the analyte molecule bound to the first reactive group 1 and does not make a linkage between them. In still other embodiments, the interaction of the affinity binder 804 of the reporter molecule 803 and the analyte molecule bound to the first reactive group 1 is not strong to make a linkage between them without cooperatively inducing by the linkage between the reporter molecule 803 and the second reactive group 2.

[00099] In some embodiments, after each iteration of contacting a solution containing reporter molecules with the supramolecular structures and subsequent incubation and identification (910), the reporter molecules are separated from the supramolecular structures via a mild denaturing step (912). In some embodiments, the mild denaturing step comprises contacting the supramolecular structures with a low concentration of urea, detergent, formamide, or a combination thereof. Accordingly, after each denaturing step (912), another solution containing reporter molecules is contacted (906) with the supramolecular structures, wherein steps 908, 910, 912 and 914 are repeated.

[000100] In some embodiments, wherein one or more supramolecular structure are identified as not having a reporter molecule linked thereto (e.g., via step 914), the reporter molecule is tuned so as to boost an affinity interaction between the reporter molecule and said supramolecular structures. For example, in some embodiments, tuning the reporter molecule comprises replacing the affinity binder with another affinity binder, and/or modifying the tail section of the reporter molecule. In some embodiments, modifying the tail section of the reporter molecule comprises increasing a sequence length of a corresponding nucleic acid. In some embodiments, modifying the tail section of the reporter molecule comprises changing a sequence of a corresponding nucleic acid. Accordingly, in some embodiments, by increasing the sequence length, the affinity binder of the reporter molecule may subsequently form a linkage with the analyte molecule of a supramolecular structure previously identified as having no interaction thereto, despite the weak affinity between the analyte molecule and affinity binder.

[000101] As depicted in (914) of FIG. 9B, after performing steps (906) to (912) a number of times, each supramolecular structure (identified via a location on a substrate or array, e.g., x, y, coordinate) is indicated (e.g., via being recorded) as being identified with a reporter molecule linked thereto (via a “1”) or identified without a reporter molecule linked thereto (“0”) with each iteration (e.g., cycle) of contacting a solution with reporter molecules. In some embodiments, each identified detection of a reporter molecule corresponds to a specific affinity binder. Accordingly, in some embodiments, for a given supramolecular structure, the total iterations (e.g., cycles) identified as having a detected reporter molecule linked thereto corresponds to specific combination of affinity binders, which are correlated to detect and identify a corresponding analyte molecule.

[000102] In another aspect, the array of supramolecular structure is configured to produce an emission signal upon exposure to electromagnetic radiation sufficient to excite the at least one signaling molecules upon binding of the affinity molecule onto the analyte molecule. In some embodiments, one or more light sensing imaging devices are configured to acquire a plurality of pixel information of the emission signal of the array, and a non-transitory computer-readable storage medium comprises machine-executable code that, upon execution by one or more computer processors.

[000103] In another embodiments, the non-transitory computer-readable medium comprises machine-executable code that, upon execution by one or more computer processors, implements a method for identifying one or more components of an array of supramolecular structure. In some embodiments, identifying one or more components of an array of supramolecular structure comprises: obtaining the array of supramolecular structure, wherein the array is configured to produce an emission signal upon exposure to electromagnetic radiation sufficient to excite the at least one signaling molecule on the array; using one or more light sensing devices configured to acquire a plurality of pixel information of the emission signal of the array, acquiring a plurality of pixel information of the array; classifying each of the plurality of pixel information into a categorical classification from among a plurality of distinct categorical classifications, thereby producing a plurality of pixel classifications; and identifying one or more components of the array of supramolecular structure based at least in part on the pixel classifications.

[000104] While preferred embodiments of the present invention have been shown and described herein, it will be obvious to those skilled in the art that such embodiments are provided by way of example only. It is not intended that the invention be limited by the specific examples provided within the specification. While the invention has been described with reference to the aforementioned specification, the descriptions and illustrations of the embodiments herein are not meant to be construed in a limiting sense. Numerous variations, changes, and substitutions will now occur to those skilled in the art without departing from the invention. Furthermore, it shall be understood that all aspects of the invention are not limited to the specific depictions, configurations or relative proportions set forth herein which depend upon a variety of conditions and variables. It should be understood that various alternatives to the embodiments of the invention described herein may be employed in practicing the invention. It is therefore contemplated that the invention shall also cover any such alternatives, modifications, variations or equivalents. It is intended that the following claims define the scope of the invention and that methods and structures within the scope of these claims and their equivalents be covered thereby.

Claims

CLAIMS WHAT IS CLAIMED IS:

1. A nucleic acid having a sequence comprising a set of segments comprising a first segment and a second segment, wherein the first segment identifies a first affinity binder to an analyte molecule and the second segment identifies a second affinity binder to the analyte molecule, wherein the first affinity binder and the second affinity binder are different; wherein each of the first affinity binder and the second affinity binder is configured to bind to the analyte molecule with a binding affinity of a dissociation constant (K_d) value, less than about 1 pM, and wherein the set of segments identifies the analyte molecule.

2. A nucleic acid having a sequence comprising a set of segments having N segments, wherein N is greater than 1, wherein each segment identifies an affinity binder in a set of affinity binders, wherein each affinity binder in the set of affinity binders is different from other affinity binders in the set of affinity binders, and wherein each affinity binder is configured to bind to an analyte molecule with a binding affinity of a dissociation constant (Kd) value, less than about 1 pM, and wherein the set of segments identifies the analyte molecule.

3. The nucleic acid of claim 2, wherein N is 3, 4, 5, 6, 7, 8, 9, or 10.

4. The nucleic acid of claim 2 wherein N is greater than 10.

5. A structure comprising the nucleic acid of claim 2 linked to a supramolecular structure.

6. The nucleic acid of any preceding claim, wherein the nucleic acid is formed by a process comprising: a. providing a supramolecular structure comprising: i. a core structure comprising a plurality of core molecules, ii. a first reactive group linked to the core structure at a first location and configured to form a linkage with the analyte molecule, and iii. a second reactive group linked to the core structure at a second location and configured to form a linkage with a first binding molecule comprising i) a first affinity binder and ii) a first tail section comprising a first barcode corresponding to the first segment of the nucleic acid; b. binding the analyte molecule to the first reactive group; c. linking i) the first affinity binder to the analyte molecule, and ii) the first binding molecule to the second reactive group via the first tail section; d. separating i) the first affinity binder from the analyte molecule, and ii) a portion of the first tail section from the first barcode, such that the first barcode remains linked to the second reactive group; e. contacting the analyte molecule with a second binding molecule comprising i) a second affinity binder and ii) a second tail section comprising a second barcode corresponding to the second segment, such that i) the second affinity binder links with the analyte molecule, and ii) the second binding molecule links to the first barcode via the second tail section; f. separating i) the second affinity binder from the analyte molecule, and ii) a portion of the second tail section from the second barcode, such that the second barcode remains linked to the first barcode, wherein the set of segments comprise the first barcode and the second barcode.

7. The process of claim 6, further comprising repeating steps (e)-(f) one or more times each with a different binding molecule, such that the set of segments comprises the first barcode, the second barcode, and at least one barcode corresponding to a binding molecule from the one or more times where steps (e)-(f) was repeated.

8. The process of any one of claims 1-7, wherein the sequence comprises a unique nucleic acid strand identified via DNA sequencing.

9. The process of any one of claims 1-8, further comprising quantifying the concentration of the analyte molecule in the sample.

10. The process of any one of claims 1-9, further comprising identifying the detected analyte molecule.

11. The process of any one of claims 6-10, wherein the analyte molecule binds with the first reactive group via a chemical bond.

12. The process of claim 11, wherein the chemical bond comprises a covalent bond.

13. The process of claim 11, wherein the chemical bond comprises a non-covalent bond.

14. The process of any one of claims 6-13, wherein the second reactive group comprises a single stranded nucleic acid of a repeating specific sequence.

15. The process of any one of claims 6-14, wherein the tail section of the first binding molecule comprises a double stranded nucleic acid comprising a first strand of a first barcode sequence and a second strand of a sequence complementary to a repeating sequence of the second reactive group.

16. The process of any one of claims 6-15, wherein the tail section of the binding molecule links with the second reactive group via hybridization of nucleic acids.

17. The process of any one of claims 6-15, wherein the tail section of the binding molecule binds to the second reactive group via formation of a phosphodiester bond between nucleic acids.

18. The process of any one of claims 6-17, wherein the tail section of each binding molecule of the one or more binding molecules further comprises one or more reactive binders configured to link the corresponding affinity binder to the corresponding barcode.

19. The process of any one of claims 6-18, prior to step (c), further comprising incubating a sample containing the analyte molecule, with the supramolecular structure for a time period.

20. The process of claim 19, wherein the time period is from about Is to about 24 hrs.

21. The process of any one of claims 6-20, wherein the barcode of each binding molecule of the one or more binding molecules is a single stranded or double stranded nucleic acid having a unique sequence.

22. The process of any one of claims 6-21, wherein the at least one of the one or more reactive binders is cleavable, so as to separate the barcode from the affinity binder.

23. The process of any one of claims 6-22, wherein the tail section of each binding molecule of the one or more binding molecules further comprises a tail reactive group configured to link with the second reactive group and/or with the barcode from a different binding molecule of the one or more binding molecules.

24. The process of any one of claims 6-23, wherein linking the first binding molecule to the second reactive group comprises ligating a first tail reactive group of the first tail section with the second reactive group.

25. The process any one of claims 6-24, further comprising, prior to step (d), further comprising contacting the supramolecular structure with a reagent solution, thereby enabling separating the affinity binder and the portion of the first tail section from the first barcode.

26. The process of claim 25, wherein, the reagent solution comprises a low concentration of detergent, urea, formamide, or any combination thereof.

27. The process of any one of claims 6-26, wherein separating the portion of the first tail section from the first barcode is via strand displacement.

28. The process of claim 27, wherein, strand displacement is via a single-stranded DNA comprising a sequence complementary to the sequence of a portion of the tail section of the binding molecule.

29. The process of any one of claims 6-28, further comprising binding the barcode with a protection binder wherein the protection binder is a single stranded nucleic acid of a sequence complementary to the sequence of the barcode.

30. The process of any one of claims 6-29, further comprising providing a sample containing the analyte molecule, wherein the sample comprises a complex biological sample and the process provides for single-molecule sensitivity thereby increasing a dynamic range and quantitative capture of a range of molecular concentrations within the complex biological sample.

31. The process of any one of claims 6-30, wherein the analyte molecule comprises a protein, a peptide, a peptide fragment, a lipid, a DNA, a RNA, an organic molecule, an inorganic molecule, complexes thereof, or any combinations thereof.

32. The process of any one of claims 6-31, wherein each supramolecular structure is a nanostructure.

33. The process of any one of claims 6-32, wherein each core structure is a nanostructure.

34. The process of any one of claims 6-33, wherein the plurality of core molecules for each core structure are arranged into a pre-defmed shape and/or have a prescribed molecular weight.

35. The process of claim 34, wherein the pre-defmed shape is configured to limit or prevent cross-reactivity with another supramolecular structure.

36. The process of any one of claims 6-35, wherein the plurality of core molecules for each core structure comprises one or more nucleic acid strands, one or more branched nucleic acids, one or more peptides, one or more small molecules, or combinations thereof.

37. The process of any one of claims 6-36, wherein each core structure independently comprises a scaffolded deoxyribonucleic acid (DNA) origami, a scaffolded ribonucleic acid (RNA) origami, a scaffolded hybrid DNA:RNA origami, a single-stranded DNA tile structure, a multi -stranded DNA tile structure, a single-stranded RNA origami, a multi -stranded RNA tile structure, hierarchically composed DNA or RNA origami with multiple scaffolds, a peptide structure, or combinations thereof.

38. The process of any one of claims 6-37, wherein the first reactive group comprises an amine, a thiol, a DBCO, a maleimide, a streptavidin, a biotin, an azide, an acrydite, a NHS-ester, a single stranded nucleic acid (RNA or DNA) of specific sequence, one or more polymers like PEG or polymerization initiators, or a combination thereof.

39. The process of any one of claims 6-38, wherein the second reactive group comprises a single stranded nucleic acid of a specific sequence.

40. The process of any one of claims 6-39, wherein the second reactive group comprises a terminal phosphate, amine, maleimide, or a combinations thereof such that it forms a chemical linkage with a single stranded nucleic acid.

41. The process of claim 40, wherein the chemical linkage comprises a phosphodiester bond.

42. The process of any one of claims 6-41, wherein the first location and the second location are pre-determined or programmable on the core structure.

43. A process for identifying an analyte molecule present in a sample, the process comprising: a. providing a supramolecular structure comprising: i. a core structure comprising a plurality of core molecules, ii. a first reactive group linked to the core structure at a first location and configured to form a linkage with an analyte molecule, and iii. a second reactive group linked to the core structure at a second location that is spaced apart from the first location by a prescribed distance, the second reactive group configured to form a linkage with one or more reporter molecules wherein each reporter molecule comprises i) an affinity binder configured to bind with the analyte molecule, ii) a tail section, and iii) at least one signaling molecules linked between the affinity binder and the tail section; b. contacting the supramolecular structure with the sample, such that the analyte molecule interacts with the first reactive group and is bound thereto; c. contacting a first solution comprising a first reporter molecule of the one or more reporter molecules with the supramolecular structure; d. detecting i) the supramolecular structure linked with the first reporter molecule via a first signaling molecule of the first reporter molecule, wherein a first affinity binder of the first reporter molecule is linked to the analyte molecule, and a first tail section of the first reporter molecule is linked to the second reactive group, or ii) the supramolecular structure not being linked to the first reporter molecule; e. when the supramolecular structure is linked with the first reporter molecule, separating the first reporter molecule from the supramolecular structure; f. contacting a second solution comprising a second reporter molecule of the one or more reporter molecules with the supramolecular structure, g. detecting i) the supramolecular structure linked with the second reporter molecule via a second signaling molecule of the second reporter molecule, wherein a second affinity binder of the second reporter molecule is linked to the analyte molecule, and a second tail section of the second reporter molecule is linked to the second reactive group, or ii) the supramolecular structure not being linked to the second reporter molecule; h. identifying the analyte molecule based on detecting the first and/or second reporter molecule being linked to the supramolecular structure.

44. The process of claim 43, when the supramolecular structure is detected not being linked to the first reporter molecule, further comprising prior to step (e), tuning the second reporter molecule such that i) the second affinity binder is different from the first affinity binder, ii) the second tail section comprises a modified first tail section to increase an affinity strength with the second reactive molecule, or iii) a combination thereof.

45. The process of any one of claims 43-44, wherein tuning the first reporter molecule comprises increasing a length of a nucleic acid sequence of the tail section nucleic acid.

46. The process of claim 45, wherein the nucleic acid sequence of the tail section nucleic acid comprises from about 2 bases to about 10 bases.

47. The process of claim 45, wherein tuning the first report molecule comprises increasing the length of a nucleic acid sequence to at most about 12 bases.

48. The process of any one of claims 43-44, wherein tuning the first reporter molecule comprises changing a nucleic acid sequence of the tail section nucleic acid.

49. The process of any one of claims 43-48, prior to step (h), repeating steps (c)-(e) one or more times each with a different reporter molecule of the one or more reporter molecules, such that the identifying the analyte molecule is based on detecting the first reporter molecule, the second molecule, and/or any combination of one or more reporter molecules for the one or more times where steps (c)-(e) is repeated.

50. The process of any one of claims 43-49, further comprising quantifying the concentration of the analyte molecule in the sample.

51. The process of any one of claims 43-50, wherein the analyte molecule interacts with the first reactive group via a chemical bond.

52. The process of claim 51, wherein the chemical bond comprises a covalent bond.

53. The process of claim 51, wherein the chemical bond comprises a non-covalent bond.

54. The process of any one of claims 43-53, prior to step (c), incubating the sample containing the analyte molecule, with the supramolecular structure for a time period.

55. The process of claim 54, wherein the time period is from about Is to about 24 hrs.

56. The process of any one of claims 43-55, wherein the tail section of each reporter molecule of the one or more reporter molecules is a single stranded or double stranded nucleic acid.

57. The process of any one of claims 43-56, wherein the signaling molecule comprises a fluorophore, a redox molecule, or a magnetically active molecule.

58. The process of any one of claims 43-57, wherein the prescribed distance is about 1 nm to about 100 nm.

59. The process of any one of claims 43-58, further comprising, prior to step (e), further comprising contacting the supramolecular structure with a reagent solution, thereby enabling separating the first reporting molecule from the supramolecular structure.

60. The process of claim 59, wherein, the reagent solution comprises a mild detergent, urea, formamide, or any combination thereof.

61. The process of any one of claims 43-60, wherein the detecting the supramolecular structure being linked with the first reporter molecule is via a imaging the supramolecular structure.

62. The process of any one of claims 43-61, further comprising the recording the detection of step (d) and (g).

63. The process of any one of claims 43-62, wherein the sample comprises a complex biological sample and the process provides for single-molecule sensitivity thereby increasing a dynamic range and quantitative capture of a range of molecular concentrations within the complex biological sample.

64. The process of any one of claims 43-63, wherein the analyte molecule comprises a protein, a peptide, a peptide fragment, a lipid, a DNA, a RNA, an organic molecule, an inorganic molecule, complexes thereof, or any combinations thereof.

65. The process of any one of claims 43-64, wherein each supramolecular structure is a nanostructure.

66. The process of any one of claims 43-65, wherein each core structure is a nanostructure.

67. The process of any one of claims 43-66, wherein the plurality of core molecules for each core structure are arranged into a pre-defmed shape and/or have a prescribed molecular weight.

68. The process of claim 67, wherein the pre-defmed shape is configured to limit or prevent cross-reactivity with another supramolecular structure.

69. The process of any one of claims 43-68, wherein the plurality of core molecules for each core structure comprises one or more nucleic acid strands, one or more branched nucleic acids, one or more peptides, one or more small molecules, or combinations thereof.

70. The process of any one of claims 43-69, wherein each core structure independently comprises a scaffolded deoxyribonucleic acid (DNA) origami, a scaffolded ribonucleic acid (RNA) origami, a scaffolded hybrid DNA:RNA origami, a single-stranded DNA tile structure, a multi -stranded DNA tile structure, a single-stranded RNA origami, a multi -stranded RNA tile structure, hierarchically composed DNA or RNA origami with multiple scaffolds, a peptide structure, or combinations thereof.

71. The process of any one of claims 43-70, wherein the first reactive group comprises an amine, a thiol, a DBCO, a maleimide, a streptavidin, a biotin, an azide, an acrydite, a NHS-ester, a single stranded nucleic acid (RNA or DNA) of specific sequence, one or more polymers like PEG or polymerization initiators, or a combination thereof.

72. The process of any one of claims 43-71, wherein the second reactive group comprises a single stranded nucleic acid of a specific sequence.

73. The process of any one of claims 6-72, wherein the supramolecular structure further comprises an identifier molecule linked thereto.

74. The process of claim 73, wherein the identifier molecule comprises DNA (e.g., of a specific sequence), RNA (e.g., of a specific sequence), fluorophore, or a combination thereof.