EP1676122A2 - Procedes pour ameliorer l'analyse de detection de particules - Google Patents

Procedes pour ameliorer l'analyse de detection de particules

Info

Publication number
EP1676122A2
EP1676122A2 EP04789395A EP04789395A EP1676122A2 EP 1676122 A2 EP1676122 A2 EP 1676122A2 EP 04789395 A EP04789395 A EP 04789395A EP 04789395 A EP04789395 A EP 04789395A EP 1676122 A2 EP1676122 A2 EP 1676122A2
Authority
EP
European Patent Office
Prior art keywords
particle
particles
electromagnetic radiation
radiation signal
analytical
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP04789395A
Other languages
German (de)
English (en)
Inventor
Robert S. Puskas
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Singulex Inc
Original Assignee
Singulex Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Singulex Inc filed Critical Singulex Inc
Publication of EP1676122A2 publication Critical patent/EP1676122A2/fr
Withdrawn legal-status Critical Current

Links

Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume, or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G01N15/14—Electro-optical investigation, e.g. flow cytometers
    • G01N15/1456—Electro-optical investigation, e.g. flow cytometers without spatial resolution of the texture or inner structure of the particle, e.g. processing of pulse signals
    • G01N15/1459—Electro-optical investigation, e.g. flow cytometers without spatial resolution of the texture or inner structure of the particle, e.g. processing of pulse signals the analysis being performed on a sample stream
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume, or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G01N15/14—Electro-optical investigation, e.g. flow cytometers
    • G01N15/1429—Electro-optical investigation, e.g. flow cytometers using an analyser being characterised by its signal processing
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume, or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G01N15/14—Electro-optical investigation, e.g. flow cytometers
    • G01N15/1425—Electro-optical investigation, e.g. flow cytometers using an analyser being characterised by its control arrangement
    • G01N15/1427—Electro-optical investigation, e.g. flow cytometers using an analyser being characterised by its control arrangement with the synchronisation of components, a time gate for operation of components, or suppression of particle coincidences
    • G01N15/1433—
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume, or surface-area of porous materials
    • G01N2015/0092—Monitoring flocculation or agglomeration
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume, or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G01N15/14—Electro-optical investigation, e.g. flow cytometers
    • G01N2015/1402—Data analysis by thresholding or gating operations performed on the acquired signals or stored data
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume, or surface-area of porous materials
    • G01N15/10—Investigating individual particles
    • G01N15/14—Electro-optical investigation, e.g. flow cytometers
    • G01N15/1434—Electro-optical investigation, e.g. flow cytometers using an analyser being characterised by its optical arrangement
    • G01N2015/1438—Using two lasers in succession

Definitions

  • the data cross-correlations will be large at values of j where the first data set from a detector (preferably photon counts above a background level) (g) resembles the data set (h) from a second detector (preferably above a background level) at some lag time (j) that corresponds to the time for specific particles to pass from the first detector to the second detector (preferably in a single molecule analytical system).
  • the lag time (j) for detection between photon detectors arrayed along the length of capillary is related to the electrophoretic velocity of a detected particle.
  • Event refers to a cross-correlated signal. Events may or may not be the result of fluorescence from a target particle. Events are considered to be of interest if they meet additional criteria known to match the characteristics of the target particle.
  • Fluid As used herein, the term “fluid” is a medium wherein particles are suspended and move. It can be gaseous, aqueous, non-aqueous, or any combination thereof. In some cases, it can have an electric field or conduct an electrical current. It may further contain salts, ions, polymers, macromolecules, or other agents that can interact with the polypeptides or polynucleotides and influence their movement.
  • Fluorescence refers to the photons of energy that are emitted as an excited fluorophore returns to its ground state. The energy of the emitted photon is usually, but not always lower, and therefore of longer wavelength, than the excitation photon.
  • Fluorescence Burst Duration As used herein, the term “fluorescence burst duration” refers to the period of time during which an emission event is detected. A synonymous term is peak width.
  • Fluorescence Intensity As used herein, the term “fluorescence intensity” refers to the total number of photons measured during a single time segment (e.g., over a millisecond and above a background level).
  • Mass tags As used herein, the term “mass tag” refers to any mass added to the target that serves to distinguish the mass tag+target from the target alone based on detection of the mass, or charge to mass ratio. A mass tag can be a label.
  • Particle As used herein, the term “particle” means an entity that can be detected, counted and/or discriminated in the current invention. Examples of particles are proteins, nucleic acids, nanospheres, microspheres, aggregates, dendrimers, organelles, chromosomes, carbohydrates, micelles, viruses, bacteria, cells, prions, and chemical entities (such as amino acids, nucleotides, lipids, sugars, toxins, venoms, drugs, reaction products and substrates).
  • ultrasensitive detection include monitoring for bioterror agents, medical application such as in the detection of drugs of abuse, biomarkers for therapeutic dosage monitoring, health status, donor matching for transplantation purposes, pregnancy, and detection of disease, pathogens, and the like, and applications in environmental, ecological, and industrial monitoring, manufacturing process monitoring and food safety.
  • Achieving the goal of single particle detection is within the scope of laser-induced detection systems; however, the lower the detection level, the more challenging it is to maximize the signal to background ratio.
  • various methods have been implemented to reduce background radiation such as using very small interrogation volumes, specific band pass filters, pulsed lasers with time-gated detection, and near-infra red emission and detection. Methods of data analysis can also be used to discriminate true signals from background.
  • particles include labels that were bound to target particles, separated from unbound labels, and interacted with an agent causing the release of the bound labels.
  • These released labels can be considered as particles, and analyzed by the methods of the current invention, thereby indirectly detecting the original target particle.
  • detection of microorganisms and cells including viruses, prokaryotic and eukaryotic cells, unicellular and multicellular organism cells, e.g., fungi, animal, mammal, or fragments thereof.
  • the methods of the invention may also be used for detecting pathogens.
  • Pathogens of interest may be, but are not limited to, viruses such as Herpesviruses, Poxviruses, Togaviruses, Flaviviruses, Picornaviruses, Orthomyxoviruses, Paramyxoviruses, Rhabdoviruses, Corona viruses, Arenaviruses, and Retroviruses.
  • viruses such as Herpesviruses, Poxviruses, Togaviruses, Flaviviruses, Picornaviruses, Orthomyxoviruses, Paramyxoviruses, Rhabdoviruses, Corona viruses, Arenaviruses, and Retroviruses.
  • bacteria including but not limited to Escherichia coli, Pseudomonas aeruginosa, Enterobacter cloacae, Staphylococcus aureus, Enterococcus faecalis, Klebsiella pneumoniae, Salmonella typhimurium, Staphylococcus epidermidis, Serratia marcescens, Mycobacterium bovis, methicillin resistant Staphylococcus aureus and Proteus vulgaris.
  • pathogens are not limited to those listed above, and one skilled in the art will know which specific species of microorganisms and parasites are of particular importance. The non-exhaustive list of these organisms and associated diseases can be found for example in U.S.
  • test sample can be pre-treated prior to use, such as preparing plasma from blood, diluting viscous fluids, or the like; methods of treatment can involve filtration, distillation, concentration, inactivation of interfering compounds, and the addition of reagents.
  • Extrinsic properties are those that are provided by a label when it is attached to the particle. Labels are applied before, after, or simultaneously with positioning the particle into the interrogation fluid. Once a particle is detectably labeled, any suitable means of detection that are known in the art can be used. Different characteristics of the electromagnetic radiation may be detected including: emission wavelength, emission intensity, burst size, burst duration and fluorescence polarization. The only proviso is that the means of detection can be used in accordance with an SMD instrument such as that provided in U.S. Patent No. 4,793,705, incorporated herein by reference in its entirety. A particle may be detectable based on any combination of intrinsic and extrinsic properties. Preferably, the means of detection is a fluorescent label.
  • a dye may be intercalating, or be noncovalently or covalently bound to a particle.
  • Dyes themselves may constitute probes such as dye probes that detect minor groove structures, cruciforms, loops or other conformational elements of particles.
  • the label may be non-fluorescent in the unbound state, but become fluorescent through changes that occur in the molecule when it binds to the target particle.
  • fluorescent markers such as fluorescent particles, fluorescent conjugated antibodies, or the like
  • the sample may be irradiated with light that is absorbed by the fluorescent particles and the emitted light measured by light measuring devices.
  • Useful light scattering tags include metals such as gold, selenium and titanium oxide, as well as nanoclusters of materials, such as ceramics or metals.
  • the exact method for attaching the bead to the particle is not critical to the practice of the invention, and a number of alternatives are known in the art.
  • the attachment is generally through interaction of the particle with a specific binding partner that is conjugated to the coating on the bead and provides a functional group for the interaction.
  • Antibodies are examples of binding partners.
  • Antibodies may be coupled to one member of a high affinity binding system, e.g., biotin, and the particles attached to the other member, e.g., avidin.
  • One may also use secondary antibodies that recognize species-specific epitopes of the primary antibodies, e.g., anti-mouse Ig, anti-rat Ig.
  • Optical tags are well known to one skilled in the art and include any entity that augments the optical properties of a target particle when bound to that particle. Examples are beads, quantum dots, or other molecules that might affect properties such as reflectivity or absorbance.
  • the extrinsic properties that render the particle detectable are provided by at least two labels of characterized photon yield.
  • the target particle is labeled with two or more labels and each label is distinct due to detected emission at one or more wavelengths that is distinguishable from the emission of the other label(s).
  • the particle is distinguished from free label by the ratio of detected emission at two or more wavelengths.
  • the particle is labeled with two or more labels and at least two of the labels emit at the same wavelength.
  • particles are distinguished based on the difference in the intensity of the detected fluorescence produced by emission from the two, three, or more labels attached to each particle.
  • the dyes have the same or overlapping excitation spectra, but possess distinguishable emission spectra.
  • dyes are chosen such that they possess substantially different emission spectra, preferably having emission maxima separated by greater than 10 nm, more preferably having emission maxima separated by greater than 25 nm, even more preferably separated by greater than 50 nm.
  • the second label may quench the fluorescence of the first label, resulting in a loss of fluorescent signal for doubly labeled particles.
  • fluorescencing/quenching pairs examples include 5' 6-FAMTM/3' Dabcyl, 5' Oregon Green ® 488-X NHS Ester/3' Dabcyl, 5' Texas Red ® -X NHS Ester/3' BlackHole QuencherTM-1 (Integrated DNA Technologies, Coralville, IA).
  • FRET fluorescence resonance energy transfer
  • excitation is transferred from the donor to the acceptor molecule without emission of a photon from the donor.
  • the donor and acceptor molecules must be in close proximity (1-10 nm).
  • the labeled particle must be distinguished from unbound label.
  • unbound label is separated from labeled particles prior to analysis.
  • the assay is a homogenous assay, and the sample, including unbound label, is analyzed by a combination of electrophoresis and single particle fluorescence detection.
  • electrophoretic conditions are chosen which provide distinct velocities for the labeled particle and the unbound label.
  • Non-specific labeling of nucleic acids generally labels all nucleic acids regardless of the particular nucleotide sequence.
  • nucleic acids include: intercalating dyes such as TOTO, ethidium bromide, and propidium iodide, ULYSIS kits for formation of coordination complexes, ARES kits for incorporation of a chemically reactive nucleotide analog to which a label can be readily attached, and incorporation of a biotin containing nucleotide analog for attachment of a streptavidin bound label. Enzymatic incorporation of labeled nucleotide analogs is another approach. Techniques to non-specifically label proteins are also well known to one skilled in the art. Several chemically reactive amino acids on the surface of proteins have been used, for example, primary amines such a lysine.
  • binding partners are agonists and antagonists for cell membrane receptors, toxins and venoms, antibodies and viral epitopes, hormones (e.g., opioid peptides and steroids) and hormone receptors, enzymes and enzyme substrates, enzymes and enzyme inhibitors, binding cofactors and target sequences, drugs and drug targets, oligonucleotides and nucleic acids, proteins and monoclonal antibodies, antigen and specific antibody, polynucleotide and complementary polynucleotide, polynucleotide and polynucleotide binding protein; biotin and avidin or streptavidin, enzyme and enzyme cofactor; and lectin and specific carbohydrate.
  • hormones e.g., opioid peptides and steroids
  • enzymes and enzyme substrates enzymes and enzyme inhibitors
  • binding cofactors and target sequences drugs and drug targets
  • oligonucleotides and nucleic acids proteins and monoclonal antibodies
  • antigen and specific antibody polynucleotide and complementary
  • Illustrative receptors that can act as a binding partner include naturally occurring receptors, e.g., thyroxine binding globulin, lectins, various proteins found on the cell surfaces (e.g., cluster of differentiation or cluster designation, or CD molecules), and the like.
  • An example is CD4, the molecule that primarily defines helper T lymphocytes.
  • a binding partner may specifically bind to related particles.
  • An example would be a peptide that binds to a family of related enzymes.
  • a sample is reacted with beads or microspheres that are coated with a binding partner that reacts with the target particle. The beads are separated from any non-bound components of the sample, and the analyzer of the invention detects the beads containing bound particles.
  • Fluorescently stained beads are particularly well suited for these methods.
  • fluorescent beads coated with oligomeric sequences will specifically bind to target complementary sequences, and after the appropriate separation steps, allow for detection of the target sequence.
  • a method for detecting particles uses a sandwich assay with monoclonal antibodies as binding partners. An antibody is linked to a surface to serve as capture antibody. The sample is added and particles having the epitope recognized by the antibody would bind to the antibody on the surface. Unbound particles are washed away leaving substantially only those that are specifically bound. The bound particle/antibody can be reacted with a detection antibody that contains a detectable label. After incubating to allow reaction between the detection antibody and particles, unbound detection antibodies are washed away.
  • the particle and detection antibody can be released from the surface and detected in the instrument of the invention. Alternatively, only the detection antibody might be released and detected, thereby indirectly detecting the particle.
  • a variation would be to employ a ligand recognized by a cell receptor.
  • the ligand is bound to the surface to capture the cells that express the specific receptor.
  • the receptor could be a surface immunoglobulin, and a labeled ligand used to label the cells. Therefore, having the ligand of interest complementary to the receptor bound to the surface, cells having the specific immunoglobulin for such ligand could be detected.
  • binding partners include any entity that can produce a detectable particle such as an enzyme that converts a substrate to a fluorescent form, or a chemical that induces fluorescence in another molecule.
  • the sample to be detected may be subjected to electrophoresis. Mobility of particles within the sample fluid varies with the properties of the particle. The velocity of movement produced by electrokinetic force is determined by the relative charge and mass of the individual particle and the fluid encasing it. Movement of a particle can be altered by the type of label that has been attached to the particle, such as a charge/mass tag. Therefore, the electrophoretic velocity of each detectably labeled particle is determined. Based on the determination of the electrophoretic velocity of each detectably labeled particle, individual particles in a sample comprising multiple particles can be distinguished. Any electrophoretic separation technique combined with an immunoassay or nucleic acid hybridization labeling technique can, in principle, be adapted for use in the context of the present invention.
  • the buffer is selected from the group consisting of Gly-Gly, bicine, tricine, 2-morpholine ethanesulfonic acid (MES), 4-morpholine propanesulfonic acid (MOPS) and 2-amino-2-methyl-1-propanol hydrochloride (AMP).
  • MES 2-morpholine ethanesulfonic acid
  • MOPS 4-morpholine propanesulfonic acid
  • AMP 2-amino-2-methyl-1-propanol hydrochloride
  • An especially preferred buffer is 2 mM Tris/borate at pH 8.1 , but Tris/glycine and Tris/HCI are also acceptable.
  • Preferred ionic strength is at least 50 mM.
  • the buffer desirably further comprises a sieving matrix for use in the embodiment of the method.
  • the sieving matrix has low fluorescence background and can specifically provide size- dependent retardation of the detectably labeled particle relative to other components in the fluid.
  • the sieving matrix can be present in any suitable concentration; from about 0.1% to about 10%) is preferred. Any suitable molecular weight can be used; from about 100,000 to about 10 million is preferred.
  • Examples of sieving matrixes include poly(ethylene oxide) (PEO), poly(vinylpyrrolidone) (PVP), linear polyacrylamide and derivatives (LPA), hydroxypropyl methylcellulose (HPMC) and hydroxyethylcellulose (HEC), all of which are soluble in water and have exceptionally low viscosity in dilute concentration (0.3% wt/vol).
  • OptophoresisTM consists of subjecting particles to various optical forces, especially moving optical gradient forces. By moving the light relative to particles, typically through a medium having some degree of viscosity, particles are separated or otherwise characterized based at least in part upon the optical force asserted against the particle and the particle's dielectric constants.
  • the light sources will be lasers, and the separations are accomplished in capillary or microchannel structures that are compatible with the instrumentation described for the current invention.
  • an SMD system described in Fig. 1 may be used.
  • an analyzer of one embodiment of the present invention is designated in its entirety by the reference numeral 10.
  • the analyzer 10 includes two electromagnetic radiation sources 12, a mirror 14, a lens 16, capillary flow cells 18, two microscope objective lenses 20, two apertures 56, two detector lenses 24, two detector filters 26, two single photon detectors 28, and a processor 30 operatively connected to the detectors.
  • the radiation sources 12 are aligned so their beams 22, 24 are reflected off a front surface of mirror 14.
  • the lens 16 focuses the beams 22, 24 into two separate interrogation volumes (e.g., interrogation volumes 38, 40 shown in Fig. 2 in the capillary flow cell 18).
  • the microscope objective lenses 20 collect light from sample particles and form images of the beams 22, 24 onto the apertures 56.
  • the apertures 56 block out scattering from walls of the capillary flow cell 18.
  • the detector lenses 24 collect the light passing through the apertures 56 and focus the light onto an active area of the detectors 28 after it passes through the detector filters 26.
  • the detector filters 26 facilitate minimizing noise signals (e.g., scattered light, ambient light) and maximizing the light signal from the particle.
  • the processor 30 processes the light signal from the particle according to the methods described herein.
  • the microscope objective lenses 20 are high- numerical aperture microscope objectives.
  • the heart of the system is the glass capillary flow cell of the apparatus 18 shown in Figure 2.
  • 3. Set a threshold level above the background level, and apply analytical filters to select signals that have electromagnetic radiation signals above the threshold and form a peak. Peaks are identified independently for data collected in each detection channels. The criteria for the analytical filters fits the criteria known to match the signals of similar particles. 4.
  • the average signal is calculated using the entire number of bins in the sample.
  • a second average is calculated where bins that contain photons 2-3 standard deviations above the original background calculation are eliminated.
  • the background level is determined from the mean noise level, or the root-mean-square noise. In other cases, a typical noise value or a statistical value is chosen. In the case of single photon counting detectors, the noise is expected to follow a Poisson distribution.
  • the detected signals are selected above a threshold prior to cross-correlating the data. A threshold value is determined to discriminate true signals (peaks, bumps, particles) from background.
  • Erroneous cross-correlation can result when photons from other particles move closely behind or ahead of the "correct" photon associated with the "correct” particle.
  • at least one analytical filter is applied to the cross- correlated data that eliminates events that fall outside the known characteristics of the target particles.
  • These filters can be based on electromagnetic characteristics such as fluorescent brightness (intensity), and the width of emission signal above the threshold value (bin number). These filters are different from those applied to the signals before cross- correlation. Events can also be restricted to a certain range of elapsed time that is evaluated or a portion of the time during which the sample is analyzed. More than one filter can be applied to a data set simultaneously.
  • the methods described herein allow particles to be enumerated as they pass individually through the interrogation volumes.
  • the concentration of the sample can be determined from the number of particles counted and the volume of sample passing though the interrogation volume in a known amount of time. In the case where the interrogation volume encompasses the entire cross-section of the sample stream, only the number of particles counted and the volume passing through a cross-section of the sample stream in a known amount of time are needed to calculate the concentration the sample.
  • the concentration of the particle can be determined by interpolating from a standard curve generated with a control sample of standard concentration.
  • the concentration of the particle can be determined by comparing the measured particles to an internal or external particle standard. Knowing the sample dilution, one can calculate the concentration of particles in the starting sample.
  • Example 4 Detection and discrimination of particles in a mixture moving at different rates using optimized cross-correlation analysis and filtering.
  • IgG and a 1.1 kb PCR product were both labeled with Alexa Fluor ® 647 according to the manufacturer's protocols for proteins and nucleic acids respectively (Molecular Probes, Inc., Eugene, Oregon).

Abstract

L'invention concerne des procédés pour améliorer l'analyse de détection de particules, selon lesquels on mesure un premier signal de rayonnement électromagnétique émis par une particule, on compare par intercorrélation le signal de rayonnement électromagnétique émis par la particule, on applique un filtre analytique aux événements d'intercorrélation, améliorant ainsi l'analyse de l'émission de la particule.
EP04789395A 2003-09-30 2004-09-30 Procedes pour ameliorer l'analyse de detection de particules Withdrawn EP1676122A2 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US50724303P 2003-09-30 2003-09-30
PCT/US2004/032244 WO2005033283A2 (fr) 2003-09-30 2004-09-30 Procedes pour ameliorer l'analyse de detection de particules

Publications (1)

Publication Number Publication Date
EP1676122A2 true EP1676122A2 (fr) 2006-07-05

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EP (1) EP1676122A2 (fr)
JP (1) JP2007533971A (fr)
WO (1) WO2005033283A2 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP2204643A4 (fr) * 2007-10-29 2017-08-02 Sysmex Corporation Appareil d'analyse cellulaire et procédé d'analyse cellulaire

Families Citing this family (34)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US9040305B2 (en) 2004-09-28 2015-05-26 Singulex, Inc. Method of analysis for determining a specific protein in blood samples using fluorescence spectrometry
US8685711B2 (en) 2004-09-28 2014-04-01 Singulex, Inc. Methods and compositions for highly sensitive detection of molecules
US7333197B2 (en) * 2004-11-17 2008-02-19 Honeywell International Inc. Raman detection based flow cytometer
US9452429B2 (en) * 2006-02-02 2016-09-27 E. I. Spectra, Llc Method for mutiplexed microfluidic bead-based immunoassay
US11237171B2 (en) 2006-02-21 2022-02-01 Trustees Of Tufts College Methods and arrays for target analyte detection and determination of target analyte concentration in solution
US8460879B2 (en) 2006-02-21 2013-06-11 The Trustees Of Tufts College Methods and arrays for target analyte detection and determination of target analyte concentration in solution
EP3156799B1 (fr) * 2006-04-04 2024-01-24 Novilux, LLC Analyseur et procédé hautement sensible de détection d'analytes
CA2648385C (fr) * 2006-04-04 2020-09-01 Singulex, Inc. Systeme et procedes hautement sensibles destines a une analyse de la troponine
JP6325022B2 (ja) * 2006-04-04 2018-05-16 シングレックス，インコーポレイテッド トロポニンの分析のための高感度のシステムおよび方法
US7838250B1 (en) 2006-04-04 2010-11-23 Singulex, Inc. Highly sensitive system and methods for analysis of troponin
WO2008105893A2 (fr) 2006-06-27 2008-09-04 Biovigilant Systems, Inc. Détection d'agent pathogène par la mesure simultanée de taille/fluorescence
WO2009029073A1 (fr) 2007-08-30 2009-03-05 The Trustees Of Tufts College Procédés pour déterminer la concentration d'un analyte dans une solution
WO2009108223A2 (fr) * 2007-11-09 2009-09-03 Biovigilant Systems, Inc. Détection d’agents pathogènes par mesure simultanée de la taille/de la fluorescence
AU2008352940B2 (en) 2007-12-19 2014-06-05 Singulex, Inc. Scanning analyzer for single molecule detection and methods of use
EP3885743A1 (fr) 2008-06-10 2021-09-29 Xtralis Technologies Ltd Détection de particules
WO2010009365A1 (fr) 2008-07-18 2010-01-21 Raindance Technologies, Inc. Bibliothèque de gouttelettes
US8222047B2 (en) 2008-09-23 2012-07-17 Quanterix Corporation Ultra-sensitive detection of molecules on single molecule arrays
US8394251B2 (en) * 2008-10-07 2013-03-12 The Board Of Trustees Of The Leland Stanford Junior University Control of chemical reactions using isotachophoresis
MY158884A (en) 2009-05-01 2016-11-30 Xtralis Technologies Ltd Particle detectors
WO2010144358A1 (fr) 2009-06-08 2010-12-16 Singulex, Inc. Groupes de biomarqueurs très sensibles
WO2011109379A1 (fr) 2010-03-01 2011-09-09 Quanterix Corporation Procédés et systèmes pour étendre la plage dynamique dans des dosages pour la détection de molécules ou de particules
US9678068B2 (en) 2010-03-01 2017-06-13 Quanterix Corporation Ultra-sensitive detection of molecules using dual detection methods
US8236574B2 (en) 2010-03-01 2012-08-07 Quanterix Corporation Ultra-sensitive detection of molecules or particles using beads or other capture objects
US8415171B2 (en) 2010-03-01 2013-04-09 Quanterix Corporation Methods and systems for extending dynamic range in assays for the detection of molecules or particles
US9952237B2 (en) 2011-01-28 2018-04-24 Quanterix Corporation Systems, devices, and methods for ultra-sensitive detection of molecules or particles
WO2012142301A2 (fr) 2011-04-12 2012-10-18 Quanterix Corporation Procédé de détermination d'un protocole de traitement et/ou d'un pronostic de rétablissement d'un patient à la suite d'un traumatisme cérébral
ES2928973T3 (es) * 2012-01-11 2022-11-23 Takeda Pharmaceuticals Co Caracterización de partículas subvisibles usando un analizador de partículas
WO2014113502A1 (fr) 2013-01-15 2014-07-24 Quanterix Corporation Détection de l'adn ou de l'arn au moyen de matrices de molécules simples et d'autres techniques
WO2014141994A1 (fr) * 2013-03-15 2014-09-18 国立大学法人 東京大学 Procédé d'analyse de particules et dispositif associé
US11397163B2 (en) 2015-09-07 2022-07-26 Max-Planck-Gesellschaft Zur Foerderung Der Wissenschaften E. V. Method and apparatus for detecting particles, like biological macromolecules or nanoparticles
CN109415722A (zh) 2016-01-29 2019-03-01 普瑞珍生物系统公司 用于核酸纯化的等速电泳
WO2018148194A1 (fr) 2017-02-07 2018-08-16 Nodexus Inc. Système microfluidique avec détection combinée électrique et optique pour le tri de particules de haute précision et procédés associés
JP7088177B2 (ja) * 2017-05-24 2022-06-21 ソニーグループ株式会社 微小粒子の吸引条件の最適化方法及び微小粒子分取装置
EP3661635A4 (fr) 2017-08-02 2021-04-21 Purigen Biosystems, Inc. Systèmes, dispositifs et procédés pour l'isotachophorèse

Family Cites Families (10)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US3826364A (en) * 1972-05-22 1974-07-30 Univ Leland Stanford Junior Particle sorting method and apparatus
DE3208919A1 (de) * 1982-03-12 1983-09-22 Gesellschaft für Strahlen- und Umweltforschung mbH, 8000 München Anordnung zur messung der fluoreszenzpolarisation
US4727020A (en) * 1985-02-25 1988-02-23 Becton, Dickinson And Company Method for analysis of subpopulations of blood cells
JPH04188045A (ja) * 1990-11-21 1992-07-06 Canon Inc 検体測定装置
JPH03128434A (ja) * 1989-10-13 1991-05-31 Canon Inc 粒子測定装置
JPH0792076A (ja) * 1993-09-24 1995-04-07 Canon Inc 粒子解析装置
JPH0792077A (ja) * 1993-09-24 1995-04-07 Canon Inc 粒子解析装置
US5528045A (en) * 1995-04-06 1996-06-18 Becton Dickinson And Company Particle analyzer with spatially split wavelength filter
JPH11258144A (ja) * 1998-03-13 1999-09-24 Sysmex Corp 粒子画像解析装置
WO2003035671A2 (fr) * 2001-10-24 2003-05-01 Singulex, Inc. Procedes destines a detecter des haplotypes genetiques par interaction avec des sondes

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See references of WO2005033283A3 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP2204643A4 (fr) * 2007-10-29 2017-08-02 Sysmex Corporation Appareil d'analyse cellulaire et procédé d'analyse cellulaire

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