WO2025166149A1

WO2025166149A1 - Microfluidic devices

Info

Publication number: WO2025166149A1
Application number: PCT/US2025/014009
Authority: WO
Inventors: Geonyoung Kim; Felicia C. LINN; Jason Hao-Sean WAN
Original assignee: Life Technologies Corp; Combinati Inc
Current assignee: Life Technologies Corp; Combinati Inc
Priority date: 2024-02-02
Filing date: 2025-01-31
Publication date: 2025-08-07
Anticipated expiration: 2026-08-02
Also published as: CN224072001U

Abstract

Disclosed are microfluidic devices. The microfluidic device comprises microfeatures disposed in the loading conduit thereof, configured to separate impurities from a fluid flowing from the loading conduit to the sample compartments. The loading conduit comprises a filtration section comprising the microfeatures, and a narrow section in fluidic communication with the filtration section. The filtration section has a larger cross-sectional area than the cross-sectional area of the narrow section, resulting in added filtration capacity for the loading conduit of the microfluidic device.

Description

MICROFLUIDIC DEVICES

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application is an International Patent Application which claims the benefit of U.S. Provisional Application No. 63/549,048 file February 2, 2024, which is incorporated herein by reference.

FILED OF INVENTION

[0002] The present disclosure concerns microfluidic devices. More specifically, the disclosure is related to a microfluidic device used for a bio-analytical test. Even more specifically, the present disclosure relates to a microfluidic device that includes features for sample filtration.

BACKGROUND OF THE INVENTION

[0003] Microfluidic devices handle fluids on a small scale. Typically, a microfluidic device operates on a sub-millimeter scale and handles micro-liters, nano-liters, or smaller quantities of fluids.

[0004] Microfluidic devices can be used for processing samples for various bio-analytical tests, which generally require high accuracy in terms of sample sizes and/or sample concentrations. However, drawbacks, including contaminations/impurities, air trapping, difficulty to operate, and high production difficulties, exist for many currently available microfluidic devices, resulting in sub-optimal performance for using these microfluidic devices.

[0005] Overall, while microfluidic devices used for bio-analytical tests exist, the need for improvements in this field persists in light of at least the aforementioned drawbacks for these microfluidic devices. SUMMARY OF THE INVENTION

[0006] A solution to at least some of the above-mentioned problems associated with microfluidic devices, especially microfluidic devices used for bio-analytical tests, has been discovered. The solution resides in a loading conduit of a microfluidic device that comprises microfeatures configured to separate impurities from a fluid being loaded into the microfluidic device, resulting in reduced impurity /contaminant level in the loaded liquid samples. This can be beneficial for limiting physical and/or chemical interference caused by these impurities during the bioanalytical tests, thereby improving the accuracy of the test results. By way of example, the microfluidic devices of the present disclosure can be used for a digital PCR (dPCR) assay, which generates low intensity signals from each sample partition. Thus, interference caused by impurities in the sample fluid can have a notable negative impact on the accuracy of the test results. The microfluidic devices disclosed herein implement microfeatures (e.g., pillar arrays) in the sample loading conduit to facilitate filtration of sample fluids, which ultimately improves the performance of the dPCR tests.

[0007] Additionally, the loading conduit of the microfluidic device of the disclosure comprises various sections that have different cross-sectional areas, thereby further increasing filtration capacity of the loading conduit and reducing impurities loaded into the sample compartments. For instance, the loading conduit can include a filtration section comprising the microfeatures for separating impurities from a fluid flowing there through, and a narrow section in fluidic communication with the filtration section. The cross-sectional area of the filtration section can be greater than that of the narrow section such that impurities including large particles are not able to flow into the narrow section, thereby separating the impurities from the fluid.

[0008] Therefore, the microfluidic devices disclosed herein provide a technical achievement over at least some of the problems associated with the currently available microfluidic devices mentioned above.

[0009] Certain embodiments are directed to microfluidic devices. In certain aspects, the microfluidic device comprises a loading conduit in fluidic communication with a plurality of sample compartments, and a plurality of microfeatures disposed within the loading conduit configured to separate impurities from a fluid flowing from the loading conduit to the sample compartments. The loading conduit comprises a filtration section, and a narrow section in fluidic communication with the filtration section. The filtration section has a larger cross-sectional area than the narrow section.

[0010] Certain embodiments are directed to microfluidic devices. In certain aspects, the microfluidic device comprises a loading conduit in fluidic communication with a plurality of sample compailments, and a plurality of pillars disposed in the loading conduit configured to separate impurities from a fluid flowing from the loading conduit to the sample compartments. The loading conduit comprises a first section, a filtration section, and a narrow section. The first section, the filtration section, and the narrow section have different cross-sectional areas, configured to enhance filtration capacity of the loading conduit. The plurality of pillars is disposed in the filtration section of the loading conduit.

[0011] Certain embodiments are directed to methods of loading a liquid comprising a biological sample into the microfluidic devices disclosed herein. In certain aspects, the method comprises drawing the liquid mixture to flow through the loading conduit into the sample compartments. The liquid in the sample compartments comprises substantially no impurities with a smallest dimension of more than 10 microns.

[0012] Certain embodiments are directed to methods of processing a biological sample. In certain aspects, the method comprises applying a plurality of pressure pulses to a liquid comprising the biological sample such that the at least some of the liquid flows through the loading conduit into the sample compartments of the microfluidic device disclosed herein, and performing PCR amplification by thermal cycling the liquid in the sample compartments.

[0013] The following includes definitions of various terms and phrases used throughout this specification.

[0014] The terms “about” or “approximately” are defined as being close to as understood by one of ordinary skill in the art. In one non-limiting embodiment the terms are defined to be within 10%, preferably, within 5%, more preferably, within 1%, and most preferably, within 0.5%. [0015] The term “substantially” and its variations are defined to include ranges within 10%, within 5%, within 1%, or within 0.5%.

[0016] Chamber” refers to a structure that enables depositing in a microfluidic device of a non-sample fluid, a fluid containing a sample, such as a biological sample, or a solution or reagent that includes a sample. Examples of structures that enable sample depositing and digitization include wells, chambers, and microchambers.

[0017] ‘Conduit” refers to a structure that enables a path of movement of a sample fluid or a non-sample fluid. Examples of structures that enable paths of fluid movement include conduits, passages, microconduits, micropassages, siphon conduits, siphon passages, and siphon apertures.

[0018] “Depth” as used with reference to microfluidic devices discussed in this specification generally refers to the distance measured from the bottom of a conduit, siphon aperture or conduit, chamber, or microchamber to the top of a side wall of the conduit, siphon aperture or conduit, chamber, or microchamber, or to the gas-permeable film or thin film that covers the conduit, siphon aperture or conduit, chamber, or microchamber.

[0019] ‘Fluid” generally refers to a liquid or a gas. A fluid does not maintain a defined shape and will flow, or move such that particles of the fluid undergo a continual change in area during an observable time frame to fill a container into which it is placed. Thus, the fluid may have any suitable viscosity that permits movement. If two or more fluids are present, each fluid may be independently selected among essentially any fluid (liquids, gases, or the like) by those of ordinary skill in the art.

[0020] As used herein, the term “biological sample” means a sample or solution containing any type of biological chemical or component and/or any target molecule of interest to a user, manufacturer, or distributor of the various embodiments of the present invention described or implied herein, as well as any sample or solution containing related chemicals or compounds used for the purpose of conducting a biological assay, experiment, or test. These biological chemicals, components, or target molecules may include, but are not limited to, DNA sequences (including cell-free DNA), RNA sequences, genes, oligonucleotides, molecules, proteins, biomarkers, cells (e.g., circulating tumor cells), or any other suitable target biomolecule. A biological sample may comprise one or more of at least one target nucleic acid sequence, at least one primer, at least one buffer, at least one nucleotide, at least one enzyme, at least one detergent, at least one blocking agent, or at least one dye, marker, and/or probe suitable for detecting a target or reference nucleic acid sequence. In various embodiments, such biological components may be used in conjunction with one or more PCR methods and systems in applications such as fetal diagnostics, multiplex dPCR, viral detection, and quantification standards, genotyping, sequencing assays, experiments, or protocols, sequencing validation, mutation detection, detection of genetically modified organisms, rare allele detection, and/or copy number variation.

[0021] ‘Microfluidic” generally refers to a device, structure, article, area, system, or chip including at least one conduit, and optionally a plurality of siphon apertures or conduits, and an array of chambers or microchambers. For example, a conduit may have a cross-sectional dimension of less than or equal to about 1 millimeter, less than or equal to about 750 microns, less than or equal to about 500 microns, less than or equal to about 250 microns, less than or equal to about 100 microns, or less. A conduit or siphon conduit or aperture may have a cross-sectional dimension of less than or equal to about 50 microns, less than or equal to about 10 microns, or less.

[0022] The term “smallest dimension”, as that term is used in the specification and/or claims means the smallest value of dimensions including overall height, overall width, overall length, and overall diameter etc. for an object, a particle, or any other single unit of impurities contained in a fluid.

[0023] The term “cross-sectional area” for each section (e.g., the loading pad, the first section, the filtration section, or the narrow section) of the loading conduit disclosed throughout the specification are defined as the cross-sectional surface that is tangential to the intended flow direction of the fluid in the loading conduit.

[0024] The terms “inhibiting” or “reducing” or “preventing” or “avoiding” or any variation of these terms, when used in the claims and/or the specification, includes any measurable decrease or complete inhibition to achieve a desired result. [0025] The term “effective,” as that term is used in the specification and/or claims, means adequate to accomplish a desired, expected, or intended result.

[0026] The use of the words “a” or “an” when used in conjunction with the term “comprising,”

“including,” “containing,” or “having” in the claims or the specification may mean “one,” but it is also consistent with the meaning of “one or more,” “at least one,” and “one or more than one.”

[0027] The words “comprising” (and any form of comprising, such as “comprise” and “comprises”), “having” (and any form of having, such as “have” and “has”), “including” (and any form of including, such as “includes” and “include”) or “containing” (and any form of containing, such as “contains” and “contain”) are inclusive or open-ended and do not exclude additional, unrecited elements or method steps.

[0028] The device, sample plate, and/or instrument of the present disclosure can “comprise,” “consist essentially of,” or “consist of” particular components, compositions, etc., disclosed throughout the specification.

[0029] Other objects, features and advantages of the embodiments will become apparent from the following figures, detailed description, and examples. It should be understood, however, that the figures, detailed description, and examples, while indicating specific embodiments, are given by way of illustration only and are not meant to be limiting. Additionally, it is contemplated that changes and modifications within the spirit and scope will become apparent to those skilled in the art from this detailed description. In further embodiments, features from specific embodiments may be combined with features from other embodiments. For example, features from one embodiment may be combined with features from any of the other embodiments. In further embodiments, additional features may be added to the specific embodiments described herein.

DESCRIPTION OF THE DRAWINGS

[0030] For a more complete understanding, reference is now made to the following descriptions taken in conjunction with the accompanying drawings, in which:

[0031] FIG. 1 is a schematic diagram of a microfluidic device, according to embodiments disclosed in the specification; [0032] FIG. 2A is a schematic diagram of a top view from the inside of the filtration section in connection with the first section and the narrow section of a loading conduit, according to embodiments disclosed in the specification;

[0033] FIGS. 2B is a schematic diagram of a front view for the cross-sectional area of the first section in connection with the filtration section of a loading conduit, according to embodiments disclosed in the specification;

[0034] FIG. 3A is a schematic diagram of a sample plate, according to embodiments disclosed in the specification;

[0035] FIG. 3B is a schematic diagram of a slide that is removably securable to a frame of a sample plate, according to embodiments disclosed in the specification;

[0036] FIG. 4 shows various configurations of microfeatures in a filtration section of a loading conduit for a microfluidic device, according to embodiments disclosed in the specification.

DETAILED DESCRIPTION OF THE INVENTION

[0037] The currently available microfluidic devices for bioanalytical tests suffer several deficiencies including contamination and air trapping, difficulty to use, and high production costs. The embodiments disclosed in the specification provide a solution to at least some of these problems. The solution is premised on a microfluidic device comprising a loading conduit in fluidic communication with a plurality of sample compartments, and a plurality of microfeatures disposed within the loading conduit configured to separate impurities from a fluid flowing from the loading conduit to the sample compartments. Hence, the microfluidic device is configured to reduce the chance of contamination in the sample compartments caused by the impurities, thus improving accuracy of bioanalytical test results using the microfluidic device. Furthermore, the loading conduit can include multiple sections with different cross-sectional areas to add filtration capacity of the loading conduit, further ensuring the accuracy of the bioanalytical tests. [0038] These and other non-limiting aspects of the present invention are discussed in further detail in the following sections.

A. Microfluidic Device

[0039] Microfluidic devices are generally designed to contain and/or process biological samples for bio-analytical tests. For conventional microfluidic devices used for bio-analytical tests, particularly digital polymerase chain reaction (dPCR) tests, contamination from impurities, for example solid particles, fibers, reagent precipitants, sample matrices etc., can have a noticeable negative impact on the accuracy of the test results, due to small sample partition sizes and limited signal intensity for each sample partition during the tests. The currently available microfluidic devices can be limited in terms of implementation of integrated filtration and/or insufficient filtration capacity. The microfluidic devices disclosed in the specification implement microfeatures, for instance pillars, in a loading conduit to filtrate a liquid containing biological samples and separate out impurities, thereby improving the purity of the samples loaded in the sample compartments and the accuracy of the test results.

[0040] With reference to FIGS. 1, a schematic diagram is shown of micro fluidic device 100. Micro fluidic device 100 includes loading conduit 110 in fluidic communication with a plurality of sample compartments 208. In some embodiments, loading conduit 110 comprises loading pad 101, first section 102, filtration section 103, and narrow section 104.

[0041] According to some embodiments, loading pad 101 is configured as an inlet for receiving a fluid (e.g., a liquid containing a biological sample) therein. A cross-sectional area of loading pad 101 can be in a range of 0.47 to 0.83 mm² and all values and ranges there between including ranges of 0.47 to 0.50 mm², 0.50 to 0.53 mm², 0.53 to 0.56 mm², 0.56 to 0.59 mm², 0.59 to 0.62 mm², 0.62 to 0.65 mm², 0.65 to 0.68 mm², 0.68 to 0.71 mm², 0.71 to 0.74 mm², 0.74 to 0.77 mm², 0.77 to 0.80 mm², and 0.80 to 0.83 mm². In some embodiments, an outlet of loading pad 101 is in fluidic communication with first section 102. Loading pad 101 has a larger cross- sectional area than first section 102. The cross-sectional area of first section 102 is about 0.010 to 0.020 mm² and all values and ranges there between including ranges of 0.010 to 0.011 mm², 0.011 to 0.012 mm², 0.012 to 0.013 mm², 0.013 to 0.014 mm², 0.014 to 0.015 mm², 0.015 to 0.016 mm², 0.016 to 0.017 mm², 0.017 to 0.018 mm², 0.018 to 0.019 mm², and 0.019 to 0.020 mm². A ratio of cross-sectional area of loading pad 101 to cross-sectional area of first section 102, in some embodiments, may be in a range of 29.3 to 83 and all ranges and values there between including ranges of 29.3 to 30, 30 to 33, 33 to 36, 36 to 39, 39 to 42, 42 to 45, 45 to 48, 48 to 51, 51 to 54, 54 to 57, 57 to 60, 60 to 63, 63 to 66, 66 to 69, 69 to 72, 72 to 75, 75 to 78, 78 to 82, and 82 to 83.

[0042] In some embodiments, an outlet of first section 102 is in fluidic communication with an inlet of filtration section 103. In certain aspects, the outlet of first section 102 and the inlet of filtration section 103 form a curved boundary (as shown in FIG. 2A). The curved boundary is configured to increase filtration capacity of loading conduit 110 compared to a straight-lined boundary by increasing cross-sectional area of loading conduit 110. In some embodiments, a depth of first section 102 (di as shown in FIG. 2B) is about 80 to 120 microns and all ranges and values there between including ranges of 80 to 84 microns, 84 to 88 microns, 88 to 92 microns, 92 to 96 microns, 96 to 100 microns, 100 to 104 microns, 104 to 108 microns, 108 to 112 microns, 112 to 116 microns, and 116 to 120 microns. A depth of filtration section 103 (df as shown in FIG. 2B) is about 10 to 20 microns and all ranges and values there between including ranges of 10 to 12 microns, 12 to 14 microns, 14 to 16 microns, 16 to 18 microns, and 18 to 20 microns. Transition between the depth of first section 102 and the depth of filtration section 103 is sloped (S i as shown in FIG. 2B) to prevent entrapment of fluid. In some embodiments, first section 102 has a length (x direction as shown FIGS. 2A and 2B) of about 0.7 to 1.5 mm, and a width (y direction as shown FIGS. 2A and 2B) of about 0.10 to 0.14 mm. Filtration section 103 can have a length (x direction as shown FIGS. 2A and 2B) of about 0.2 to 0.25 mm and a width (y direction as shown FIGS. 2A and 2B) of about 0.10 to 0.14 mm.

[0043] In some embodiments, a plurality of microfeatures is disposed within filtration section 103 of loading conduit 110. The microfeatures are configured to separate impurities from a fluid flowing from loading conduit 110 to sample compartments 208. Exemplary microfeatures can include pillars, parallel channels, debris diverting pillars, and any combination thereof. Exemplary impurities can include particles, fibers, reagent precipitates, sample matrices, and any combination thereof. In some instances, the impurities can have a smallest dimension of 10 to 50 microns and all ranges and values there between including ranges of 10 to 15 microns, 15 to 20 microns, 20 to 25 microns, 25 to 30 microns, 30 to 35 microns, 35 to 40 microns, 40 to 45 microns, and 45 to 50 microns. [0044] In some embodiments, the microfeatures include pillars 105. In certain aspects, pillars 105 are disposed next to an inlet of narrow section 104. According to embodiments, pillars 105 arc further configured to limit emulsion formation and/or air trapping when the fluid flows through filtration section 103 towards narrow section 104. Each of pillars 105 can have a cross-sectional area (cross-sectional area that is tangential to the height of the pillar) substantially in a circular shape, a triangular shape, a rectangular shape, a square shape, a polygonal shape, or any combination thereof.

[0045] According to some embodiments, pillars 105 are arranged in at least one row. In some instances, pillars 105 in filtration section 103 are arranged in a staggered pattern comprising at least 3 rows. In some aspects, pillars 105 are positioned in a staggered pattern to minimize entrapment of stagnant fluid in filtration section 103 and/or facilitate directional changes in fluid flow path to increase chances of capturing impurities among pillars 105. In some embodiments, each of pillars 105 can have a substantially circular top surface and a substantially circular bottom surface. In some aspects, the bottom surface is larger than the top surface of each of pillars 105. A pillar of pillars 105 can be tapered toward the top thereof. According to embodiments, the top surface of each of pillars 105 can have a diameter of 0.018 to 0.022 mm and all ranges and values there between including ranges of 0.018 to 0.019 mm, 0.019 to 0.020 mm, 0.020 to 0.021 mm, and 0.021 to 0.022 mm. The bottom surface of each of pillars 105 can have a diameter of 0.020 to 0.028 mm and all ranges and values there between including 0.020 to 0.021 mm, 0.021 to 0.022 mm, 0.022 to 0.023 mm, 0.023 to 0.024 mm, 0.024 to 0.025 mm, 0.025 to 0.026 mm, 0.026 to 0.027 mm, and 0.027 to 0.028. A height of at least one of pillars 105 can be in a range of 0.012 to 0.016 mm and all ranges and values there between including ranges of 0.012 to 0.013 mm, 0.013 to 0.014 mm, 0.014 to 0.015 mm, and 0.015 to 0.016 mm. In some embodiments, pillars 105 comprise a cyclic olefin polymer, a cyclic olefin copolymer, or any combination thereof.

[0046] According to some embodiments, pillars 105 are arranged in 3 rows with first row containing 4 pillars, second row containing 3 pillars, and third row containing 4 pillars (rows are numbered from left to right as shown in FIG. 2A). In some embodiments, pillars 105 are positioned such that a minimum distance between two adjacent pillars is about 6 to 10 microns and all ranges and values there between including ranges of 6 to 7 microns, 7 to 8 microns, 8 to 9 microns, and 9 to 10 microns. According to embodiments, a pillar (105a or 105b shown in FIG. 2A) is disposed at each corner of an end of filtration section 103 that connects with narrow section 104 (the end that is proximal to narrow section 104). In certain aspects, pillars 105a and 105b are configured to further limit emulsion formation in the fluid flowing through filtration section 103. In embodiments, an outlet of filtration section 103 is in fluidic communication with narrow section 104. In some embodiments, the cross-sectional area of filtration section 103 is in a range of 0.0010 to 0.0025 mm² and all ranges and values there between including ranges of 0.0010 to 0.0013 mm², 0.0013 to 0.0016 mm², 0.0016 to 0.0019 mm², 0.0019 to 0.0022 mm², and 0.0022 to 0.0025 mm². The cross-sectional area of narrow section 104 is in a range of 0.00025 to 0.00046 mm², and all ranges and values there between including ranges of 0.00025 to 0.00028 mm², 0.00028 to 0.00031 mm², 0.00031 to 0.00034 mm², 0.00034 to 0.00037 mm², 0.00037 to 0.00040 mm², 0.00040 to 0.00043 mm², and 0.00043 to 0.00046 mm². A ratio of the cross-sectional area of filtration section 103 to the cross-sectional area of narrow section 104 can be in a range of 4 to 5.4 and all ranges and values there between. Narrow section 104 can have a depth of 12 to 16 microns and all ranges and values there between including ranges of 12 to 13 microns, 13 to 14 microns, 14 to 15 microns, and 15 to 16 microns. In some embodiments, pillars 105 of substantially circular top and bottom surfaces are configured to break up impurities into impurities of smaller sizes compared to pillars with other shapes of top and bottom surfaces, thereby preventing large sized impurities clogging loading conduit.

[0047] In certain aspects, sample compartments 208 of microfluidic device 100 is configured to contain partitions of a biological sample for a bioanalytical test. Sample compartments 208 can include any structures that can divide a fluid into a plurality of partitions. Exemplary sample compartments can include microchambers, through-holes, microwells, pockets, and any combination thereof.

[0048] In some embodiments, microfluidic device 100 is configured to process biological samples and/or reagents for polymerase chain reaction (PCR). In certain aspects, microfluidic device 100 is configured to contain biological samples and/or reagents for digital PCR (dPCR). Each of sample compartments 208 can have a volume of 100 to 1000 pico liters. In certain aspects, each of sample compartments 208 has a volume of 477 to 583 pico liters and all values and ranges there between including ranges of 477 to 487 pico liters, 487 to 497 pico liters, 497 to 507 pico liters, 507 to 517 pico liters, 517 to 527 pico liters, 527 to 537 pico liters, 537 to 547 pico liters, 547 to 557 pico liters, 557 to 567 pico liters, 567 to 577 pico liters, and 577 to 583 pico liters.

[0049] In some instances, sample compartments 208 include microchambers with a depth of at least about 100 microns. A ratio of a depth of the microchamber to a minimum distance between the microchamber and an adjacent microchamber can be at least about 3:1, or at least about 5:1. In certain aspects, a microchamber of sample compartments 208 comprises a substantially rectangular three-dimensional shape comprising four substantially rectangular side walls. Two adjacent sidewalls of the microchamber can be joined by a curved corner. The curved comer can have a radius of at least about 10 microns. In embodiments, microfluidic device 100 includes at least 8000 sample compartments or at least 10,000 sample compartments. In some instances, microfluidic device 100 comprises at least 20,000 sample compailments. According to embodiments, a total volume of sample compartments 208 is greater than a volume of loading conduit 110.

[0050] According to some embodiments, microfluidic device 100 further comprises a plurality of terminating chambers 307 in fluidic communication with loading conduit 110 and sample compartments 208. In some embodiments, terminating chambers 307 are configured to contain overflow, and/or residual volume of the liquid input from loading conduit 110 and/or sample compailments 208. In embodiments, loading conduit 110 can be connected to, and in fluidic communication with, a network of a plurality of branch conduits such that the input fluid can flow from loading conduit 110 to the plurality branch conduits. In some instances, microfluidic device 100 further comprises a plurality of siphon conduits, and a siphon conduit of the plurality of siphon conduits is fluidically coupled to a branch conduit of the plurality of branch conduits to a sample compartment of the plurality of sample compartments 208. In embodiments, microfluidic device 100 is configured such that an input fluid is received into loading conduit 110, and sequentially flows through a branch conduit and a siphon conduit into a sample compartment of the plurality of sample compartments 208.

B. Sample Plate

[0051] In embodiments, there are provided sample plates that are capable of improving sample quality and accuracy for bioanalytical tests compared to conventional sample plates. The sample plate can include a plurality of microfluidic devices (e.g., microfluidic device 100) as disclosed and shown in FIGS. 1, 2A and 2B.

[0052] In certain embodiments, as shown in FIG. 3A, sample plate 300 comprises frame 311 configured to secure one or more slides 312 thereon. As shown in FIG. 3B, more than two microfluidic devices (e.g., microfluidic devices 100, 100a, 100b, 100c, and lOOd) discussed above (e.g., microfluidic device 100 as shown in FIG. 1) are disposed or formed on, and/or integrated in slide 312. In some instances, slide 312 may contain 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 microfluidic devices. In embodiments, frame 311 may be substantially rectangular. Frame 311 may be configured to secure 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 slides (e.g., slide 312). According to embodiments, frame 311 can comprise a plurality of ports (e.g., port 315). Each loading pad (e.g., loading pad 101 shown in FIG. 1 and loading pads lOla-lOld shown in FIG. 3B) can correspond to and/or be in fluidic communication with a port of frame 311. The ports (e.g., port 315) may be configured to vent gas from microfluidic devices on the slides. The ports (e.g., port 315) may be further configured to receive a fluid (e.g., a biological sample) into the microfluidic devices disposed on and/or disposed in the slides (e.g., slide 312). According to some embodiments, the plurality of ports is disposed and/or formed on multiple strips (e.g., strip 316) of frame 311. According to some embodiments, each strip is positioned such that each port fits over a loading pad (e.g., loading pad 100, and lOla-lOld) of the microfluidic devices when one or more of the slides (e.g., slide 312) are secured on frame 311. In some embodiments, the strips (e.g., strip 316) may be disposed across a width or a length of frame 311.

[0053] According to embodiments, frame 311 may include a plurality of gaskets (e.g., gasket 314). Each of the gaskets is configured to be removably secured over a port (e.g., port 315). In some aspects, each gaskets comprises a cap-like structure disposed over the ports. In embodiments, the gaskets are configured to seal the ports (e.g., port 316) from outside environment. In some embodiments, the gaskets (e.g., gasket 314) are configured to removably secured to a pressure manifold. The pressure manifold, in some embodiments, is configured to apply pressure to the microfluidic devices through the gaskets. The pressure manifold, in some embodiments, is further configured to load the microfluidic devices through the gaskets. [0054] In some embodiments, sample plate 300 comprises a thin film (e.g., thin film 313) applied over the one or more microfluidic devices (e.g., microfluidic devices 100, lOOa-lOOd) configured to form a cover for sample compartments 208. The thin film can be further configured to form a cover for at least a portion of loading conduits 110, HOa-llOd. In embodiments, the thin film (e.g., thin film 313) is gas impermeable at lower pressures, but allows for out-gassing through the thin film when pressure is applied, and is thus at least partially gas permeable under pressure. In some embodiments of the invention, the gas-permeable film is not gas permeable at atmospheric pressure, but is gas-permeable at a pressure that is higher than atmospheric pressure. In some embodiments, the thin film is gas-permeable but not liquid-permeable at one or more selected pressures above atmospheric pressure. In some embodiments of the present invention, the thin film is approximately 80 microns in thickness and composed of a cyclic olefin polymer. One suitable thin film used in embodiments of the present invention is a semi-gas permeable film TOP AS® COC 6013. In other embodiments of the invention, semi-gas permeable films having a thickness of 60, 70, 80, 90, 100 microns or any range within those thicknesses may be used.

C. Method of Using Microfluidic Device

[0055] In embodiments, there are provided methods of loading a liquid comprising a biological sample into the microfluidic device disclosed above (e.g., microfluidic device 100, lOOa-lOOd). The method of loading a liquid comprising a biological sample can include drawing the liquid to flow through the loading conduit (e.g., loading conduit 110) into sample compartments 208. The liquid in the sample compartments may comprise substantially no impurities with a smallest dimension of more than 10 microns.

[0056] In embodiments, the liquid comprises reagents for conducting a PCR process. The liquid can further comprise an oil, water, or any combination thereof. The oil can include silicon oil. According to embodiments, high pressure may be applied to compress the air in the microfluidic device including the loading conduits, siphon conduits, sample compartments (e.g., microchambers), and terminating chambers, which draws the liquid into the sample compartments. The amount of fluid being drawn in should roughly equal to the air being compressed according to the ideal gas law. Since the volume of the loading conduits (having at least about 10 microns depth and at least about 10 microns width) is smaller than the volume of the sample compartments (having at least about 100 microns depth), all the loading conduits should be filled with the sample fluid upon this action, meaning most of the compressed air will stay in the sample compartments and the terminating chambers. Compressed air will continue to escape through the thin film, drawing more fluid comprising the biological sample into the loading conduits, siphon conduits and into the sample compartments (e.g., microchambers). Non-sample containing fluid overlaid on top of the sample fluid will later be drawn into the microfluidic device while the fluid comprising the biological sample continues to replace the space occupied by the air, which continues to escape through the film.

[0057] In the step of drawing the liquid comprising the biological sample to flow through the loading conduit, a series of one or more pressure pulses may be applied to the loading pad of a microfluidic device. The pressure pulse may comprise applying a high pressure for a first predetermined time period, e.g., a short time interval, followed immediately by applying a low pressure for a second predetermined time period, e.g., a short time interval. In one embodiment of the invention, the pulsing starts at the beginning (where there is no fluid in the array since the material is hydrophobic) for 1 minute (6 cycles of 75 Psi / 10 Psi for 5 seconds I 5 seconds). In other words, a higher-pressure pulse may be applied at 75 psi for a short time interval, e.g., 5 seconds, followed immediately by a lower pressure pulse at 10 psi for a short time interval, e.g., 5 seconds, followed immediately again by the higher-pressure pulse at 75 psi for 5 seconds. The higher-pressure pulse followed by the lower pressure pulse may thus repeatedly be applied for successive 5 second time intervals for 6 cycles. In other embodiments of the invention, a higher or lower number of cycles may be contemplated, e.g., 5, 10, 12, 20, or more cycles. Different higher- pressure pulses or lower-pressure pulses may also be applied, e.g., less than or greater than 10 psi for the lower-pressure pulse, or greater than or less than 75 psi for the higher-pressure pulses. According to some embodiments, a high-pressure may be continuously applied to the loading pad for a prolonged period (e.g., more than 1 minutes, or more than 5 minutes, or sometimes more than 20 minutes) to further degas the sample compartments (e.g., microchambers).

[0058] According to embodiments, the microfluidic device in the method is used for preparing samples for a digital PCR test, and each sample compartment may contain either zero, one, or more than one copies of target molecule(s). The method can further comprise performing PCR amplification by thermal cycling the liquid in the sample compartments. The method further comprises capturing images of the sample compartments of the microfluidic device and determining a number of sample compartments within which PCR amplification has been successfully achieved based on the images of the sample compartments. Prior to thermal cycling, the liquid (sample partitions) in the sample compailments contains substantially no impurities with a smallest dimension greater than 10 microns due to filtration capacity provided by the loading conduit (the microfeatures (such as pillars) in filtration section and/or various cross-sectional area sizes for each section), thereby reducing negative impact of impurities in input sample liquid on dPCR test results.

[0059] As part of the disclosure, a specific example is included below. The example is for illustrative purposes only and is not intended to limit the invention. Those of ordinary skill in the art will readily recognize parameters that can be changed or modified to yield essentially the same results.

Example

(Efficiency for Various Configurations of Microfeatures)

[0060] Eight microfeature configurations (A-E and Control as shown in FIG. 4) in the filtration section of the loading conduit were tested for filtration efficiency. A liquid with 10 micron beads at a concentration of 450 to 500 beads (fluorescent under fluorescent microscope) per test was used to load the microfluidic devices discussed above. The filtration section and the microchambers (sample compartments) were observed under microscope to determine the filtration efficiency.

[0061] Beads were observed in the filtration sections of configurations A, B, C, D, E, F and G. There were no beads observed in the filtration section of the Control. Additionally, air traps were observed in configurations B and C. Emulsion risk, manufacturability risk, and functional risk were further evaluated for configurations A, E, and G. As configuration G (only one row of pillars) had high functional risk for lack of redundancy as back up, configuration G was not selected. Configuration E included square pillar array as the microfeature for filtration, but the manufacturability risk for the square pillar array was high compared to round pillars. Therefore, configuration A was selected. Further improvements were needed for configuration A to arrive at the final design for the filtration section of the loading conduit of the microfluidic device. [0062] In the context of the specification, at least the following embodiments are described. Embodiment 1 is a microfluidic device comprising a loading conduit in fluidic communication with a plurality of sample compartments; and a plurality of microfcaturcs disposed within the loading conduit configured to separate impurities from a fluid flowing from the loading conduit to the sample compartments, wherein the loading conduit comprises a filtration section, and a narrow section in fluidic communication with the filtration section; wherein the filtration section has a larger cross-sectional area than the narrow section. Embodiment 2 is the microfluidic device of embodiment 1, wherein the loading conduit further comprises a loading pad configured to receive the fluid therein, and a first section in fluidic communication with an outlet of the loading pad, and wherein the loading pad has a larger cross-sectional area than the first section. Embodiment 3 is the microfluidic device of embodiment 2, wherein the cross-sectional area of the loading pad is in a range of 0.47 to 0.83 mm², and the cross-sectional area of the first section is in a range of 0.01 to 0.016 mm². Embodiment 4 is the microfluidic device of embodiment 2 or 3, wherein the cross- sectional area of the filtration section is in a range of 0.0010 to 0.0025 mm² and the cross-sectional area of the narrow section is in a range of 0.00025 to 0.00046 mm². Embodiment 5 is the microfluidic device of any of embodiments 2 to 4, wherein an outlet of the first section is in fluidic communication with an inlet of the filtration section. Embodiment 6 is the microfluidic device of embodiment 5, wherein the outlet of the first section and the inlet of the filtration section forms a curved boundary configured to increase filtration capacity of the loading conduit. Embodiment 7 is the microfluidic device of any of embodiments 1 to 6, wherein the microfeatures comprise pillars, debris diverting pillars, parallel channels, or a combination thereof. Embodiment 8 is the microfluidic device of embodiment 7, wherein the pillars arc disposed in the filtration section next to an inlet of the narrow section of the loading conduit. Embodiment 9 is the microfluidic device of any of embodiments 7 and 8, wherein the pillars are further configured to limit emulsion formation and air trap when the fluid flows through the filtration section. Embodiment 10 is the microfluidic device of any of embodiments 7 to 9, wherein each of the pillars has a cross-sectional area in a shape of circular, triangular, rectangular, square, polygon, or a combination thereof. Embodiment 11 is the microfluidic device of any of embodiments 7 to 10, wherein the pillars in the filtration section are arranged in at least one row. Embodiment 12 is the microfluidic device of any of embodiments 7 to 11, wherein the pillars in the filtration section are arranged in a staggered pattern comprising at least 3 rows. Embodiment 13 is the microfluidic device of any of embodiments 7 to 12, wherein the pillars are positioned such that a minimum distance between two adjacent pillars is about 6 to 10 microns. Embodiment 14 is the microfluidic device of any of embodiments 7 to 13, wherein a pillar is disposed at each corner of an end of the filtration section that connects with the narrow section. Embodiment 15 is the microfluidic device of embodiment 14, wherein the pillar at each comer of said end of the filtration section is configured to limit emulsion formation of the liquid flowing through the filtration section. Embodiment 16 is the microfluidic device of any of embodiments 7 to 15, wherein each pillar of the filtration section has a round cross-sectional area with a diameter of 20 to 24 microns. Embodiment 17 is the microfluidic device of any of embodiments 7 to 16, wherein the pillars comprise Cyclic olefin polymer , Cyclic olefin copolymer, or a combination thereof. Embodiment 18 is the microfluidic device of any of embodiments 1 to 17, wherein each of the sample compartments has a volume of about 477 to 583 pico liters. Embodiment 19 is the microfluidic device of any of embodiments 1 to 18, wherein the microfluidic device is configured to contain samples and reagents for polymerase chain reaction (PCR). Embodiment 20 is the microfluidic device of embodiment 19, wherein the microfluidic device is configured to contain samples and reagents for digital PCR (dPCR). Embodiment 21 is the microfluidic device of embodiment 20, wherein the loading conduit is in fluidic communication with at least 10,000 sample compartments.

[0063] Embodiment 22 is a microfluidic device for processing a biological sample. The microfluidic device comprises a loading conduit in fluidic communication with a plurality of sample compartments; and a plurality of pillars disposed in the loading conduit, configured to separate impurities from a fluid flowing from the loading conduit to the sample compartments; wherein the loading conduit comprises a first section, a filtration section, and a narrow section, and wherein the first section, the filtration section, and the narrow section have different cross- sectional areas, configured to enhance filtration capacity of the loading conduit. Embodiment 23 is the microfluidic device of embodiment 22, wherein the loading conduit further comprises a loading pad configured to receive a liquid into the loading conduit, and the liquid sequentially flows through the loading pad, the first section, the filtration section, and the narrow section to the sample compartments. Embodiment 24 is the microfluidic device of embodiment 23, wherein the cross-sectional area of the loading pad is in a range of 0.47 to 0.83 mm², and the cross-sectional area of the first section is in a range of 0.010 to 0.016 mm². Embodiment 25 is the microfluidic device of any of embodiments 22 and 24, wherein the cross-sectional area of the filtration section is in a range of 0.0010 to 0.0025 mm² and the cross-sectional area of the narrow section is in a range of 0.00025 to 0.00046 mm². Embodiment 26 is the microfluidic device of any of embodiments 22 to 25, wherein the pillars arc cylindrical with a cross-sectional diameter of 20 to 24 microns. Embodiment 27 is the microfluidic device of any of embodiments 22 to 26, wherein the pillars are disposed in a staggered pattern to form an array. Embodiment 28 is the microfluidic device of any of embodiments 22 to 27, wherein the pillars are disposed in the filtration section. Embodiment 29 is the microfluidic device of embodiment 28, wherein the pillars are disposed at an end of the filtration section that is proximal to the narrow section. Embodiment 30 is the microfluidic device of embodiment 29, wherein a pillar is disposed at each comer of the end of the filtration section that is proximal to the narrow section, configured to limit formation of emulsion. Embodiment 31 is the microfluidic device of any of embodiments 22 to 30, wherein the pillars are disposed in an array comprising at least one row. Embodiment 32 is the microfluidic device of embodiment 31, wherein the array comprises 3 rows with a first row including 4 pillars, a second row including 3 pillars, and a third row including 4 pillars. Embodiment 33 is the microfluidic device of any of embodiments 22 to 32, wherein the impurities include particles, fibers, reagent precipitates, sample matrices, or a combination thereof. Embodiment 34 is the microfluidic device of any of embodiments 22 to 33, wherein the impurities include particles with a smallest dimension of 10 to 50 microns. Embodiment 35 is the micro fluidic device of any of embodiments 22 to 34, wherein the fluid includes an oil, water, a nuclei acid, or a combination thereof. Embodiment 36 is the microfluidic device of any of embodiments 22 to 35, wherein the sample compartments each have a volume of 477 to 583 pico liters. Embodiment 37 is the microfluidic device of any of embodiments 22 to 36, wherein microfluidic device comprises at least 10,000 sample compartments. Embodiment 38 is the microfluidic device of any of embodiments 22 to 37, wherein microfluidic device comprises a plurality of terminating chambers in fluidic communication with the loading conduit and the sample compartments. Embodiment 39 is the microfluidic device of any of embodiments 22 to 38, wherein the terminating chambers are configured to receive overflown or residual volume of the fluid. Embodiment 40 is the microfluidic device of any of embodiments 22 to 39, further comprising a plurality of siphon conduits, a siphon conduit of the plurality of siphon conduits in fluidic communication with the loading conduit and a sample compartment. Embodiment 41 is the microfluidic device of embodiment 40, wherein the siphon conduit is configured to enable fluidic communication between the loading conduit and the sample compartments. Embodiment 42 is the microfluidic device of any of embodiments 22 to 41, further comprising a thin film applied to the microfluidic device, configured to form a cover for the sample compartments. Embodiment 43 is the microfluidic device of embodiment 42, wherein the thin film applied to the microfluidic device is further configured to form a cover for at least a portion of the loading conduit. Embodiment 44 is the microfluidic device of any of embodiments 42 and 43, wherein the thin film comprises a gas- permeable material. Embodiment 45 is the microfluidic device of any of embodiments 22 to 44, wherein a ratio of a depth of the sample compartment to a minimum distance between two adjacent sample compartments is 3.96 to 3.09. Embodiment 46 is the microfluidic device of any of embodiments 22 to 45, wherein the microfluidic device is configured to process samples for polymerase chain reaction (PCR). Embodiment 47 is the microfluidic device of any of embodiments 22 to 45, wherein the microfluidic device is configured to process samples for digital PCR (dPCR).

[0064] Embodiment 48 is a method of loading a liquid comprising a biological sample into the microfluidic device of any of embodiments 22 to 47. The method comprises drawing the liquid to flow through the loading conduit into the sample compartments; wherein the liquid in the sample compartments comprises substantially no impurities with a size of more than 10 microns. Embodiment 49 is the method of embodiment 48, wherein the liquid further comprises reagents for a PCR process. Embodiment 50 is the method of embodiment 49, wherein the liquid further comprises an oil, an aqueous solution, a nucleic acid, or combinations thereof.

[0065] Embodiment 51 is a method of processing a biological sample. The method comprises applying a plurality of pressure pulses to a liquid mixture comprising the biological sample such that the at least some liquid mixture flows through the loading conduit into the sample compartments of the microfluidic device of any of embodiments 22 to 47 ; and performing PCR amplification by thermal cycling the liquid mixture in the sample compartments. Embodiment 52 is the method of embodiment 51, wherein the liquid mixture in the sample compartments comprises substantially no impurities with a smallest dimension over 10 microns. Embodiment 53 is the method of any of embodiments 51 and 52, further comprising capturing images of the sample compartments of the microfluidic device. Embodiment 54 is the method of embodiment 53, further comprising determining a number of sample compartments within which PCR amplification has been successfully achieved based on the images of the sample compartments of the microfluidic device.

[0066] Although embodiments of the present application and their advantages have been described in detail, it should be understood that various changes, substitutions and alterations can be made herein without departing from the spirit and scope of the embodiments as defined by the appended claims. Moreover, the scope of the present application is not intended to be limited to the particular embodiments of the system, instrument, machine, manufacture, composition of matter, means, methods, and/or steps described in the specification. As one of ordinary skill in the art will readily appreciate from the above disclosure, system, instrument, processes, machines, manufacture, compositions of matter, means, methods, or steps, presently existing or later to be developed that perform substantially the same function or achieve substantially the same result as the corresponding embodiments described herein may be utilized. Accordingly, the appended claims are intended to include within their scope such processes, machines, manufacture, compositions of matter, means, methods, or steps.

Claims

CLAIMS What is claimed is:

1. A microfluidic device comprising: a loading conduit in fluidic communication with a plurality of sample compartments; and a plurality of microfeatures disposed within the loading conduit configured to separate impurities from a fluid flowing from the loading conduit to the sample compartments; wherein the loading conduit comprises a filtration section, and a narrow section in fluidic communication with the filtration section; wherein the filtration section has a larger cross-sectional area than the narrow section.

2. The microfluidic device of claim 1, wherein the loading conduit further comprises a loading pad configured to receive the fluid therein, and a first section in fluidic communication with an outlet of the loading pad, and wherein the loading pad has a larger cross-sectional area than the first section.

3. The microfluidic device of claim 2, wherein the cross-sectional area of the loading pad is in a range of 0.47 to 0.83 mm², and the cross-sectional area of the first section is in a range of 0.01 to 0.016 mm².

4. The microfluidic device of any of claims 2 and 3, wherein the cross-sectional area of the filtration section is in a range of 0.0010 to 0.0025 mm² and the cross-sectional area of the narrow section is in a range of 0.00025 to 0.00046 mm².

5. The microfluidic device of any of claims 2 to 4, wherein an outlet of the first section is in fluidic communication with an inlet of the filtration section.

6. The microfluidic device of claim 5, wherein the outlet of the first section and the inlet of the filtration section forms a curved boundary configured to increase filtration capacity of the loading conduit.

7. The microfluidic device of any of claims 1 to 6, wherein the microfeatures comprise pillars, debris diverting pillars, parallel channels, or a combination thereof.

8. The microfluidic device of claim 7, wherein the pillars are disposed in the filtration section next to an inlet of the narrow section of the loading conduit.

9. The microfluidic device of claims 7 and 8, wherein the pillars are further configured to limit emulsion formation and air trap when the fluid flows through the filtration section.

10. The microfluidic device of any of claims 7 to 9, wherein each of the pillars has a cross- sectional area in a shape of circular, triangular, rectangular, square, polygon, or a combination thereof.

11. The microfluidic device of any of claims 7 to 10, wherein the pillars in the filtration section are arranged in at least one row.

12. The microfluidic device of any of claims 7 to 11, wherein the pillars in the filtration section are arranged in a staggered pattern comprising at least 3 rows.

13. The microfluidic device of any of claims 7 to 12, wherein the pillars are positioned such that a minimum distance between two adjacent pillar s is about 6 to 10 microns.

14. The microfluidic device of any of claims 7 to 13, wherein a pillar is disposed at each comer of an end of the filtration section that connects with the narrow section.

15. The microfluidic device of claim 14, wherein the pillar at each corner of said end of the filtration section is configured to limit emulsion formation of the liquid flowing through the filtration section.

16. The microfluidic device of any of claims 7 to 15, wherein each pillar of the filtration section has a round cross-sectional area with a diameter of 20 to 24 microns.

17. The microfluidic device of any of claims 7 to 16, wherein the pillars comprise Cyclic olefin polymer , Cyclic olefin copolymer, or a combination thereof.

18. The microfluidic device of any of claims 1 to 17, wherein each of the sample compartments has a volume of about - 7 to 583 pico liters.

19. The microfluidic device of any of claims 1 to 18, wherein the microfluidic device is configured to contain samples and reagents for polymerase chain reaction (PCR).

20. The microfluidic device of claim 19, wherein the microfluidic device is configured to contain samples and reagents for digital PCR (dPCR).

21. The microfluidic device of claim 20, wherein the loading conduit is in fluidic communication with at least 10,000 sample compartments.

22. A microfluidic device for processing a biological sample, the microfluidic device comprising: a loading conduit in fluidic communication with a plurality of sample compartments; and a plurality of pillars disposed in the loading conduit, configured to separate impurities from a fluid flowing from the loading conduit to the sample compartments; wherein the loading conduit comprises a first section, a filtration section, and a narrow section, and wherein the first section, the filtration section, and the narrow section have different cross-sectional areas, configured to enhance filtration capacity of the loading conduit.

23. The microfluidic device of claim 22, wherein the loading conduit further comprises a loading pad configured to receive a liquid into the loading conduit, and the liquid sequentially flows through the loading pad, the first section, the filtration section, and the narrow section to the sample compartments.

24. The microfluidic device of claim 23, wherein the cross-sectional area of the loading pad is in a range of 0.47 to 0.83 mm², and the cross-sectional area of the first section is in a range of 0.010 to 0.016 mm².

25. The microfluidic device of any of claims 22 and 23, wherein the cross-sectional area of the filtration section is in a range of 0.0010 to 0.0025 mm² and the cross-sectional area of the narrow section is in a range of 0.00025 to 0.00046 mm².

26. The microfluidic device of any of claims 22 to 25, wherein the pillars are cylindrical with a cross-sectional diameter of 20 to 24 microns.

27. The microfluidic device of any of claims 22 to 26, wherein the pillars are disposed in a staggered pattern to form an array.

28. The microfluidic device of any of claims 22 to 27, wherein the pillars are disposed in the filtration section.

29. The microfluidic device of claim 28, wherein the pillars are disposed at an end of the filtration section that is proximal to the narrow section.

30. The microfluidic device of claim 29, wherein a pillar is disposed at each corner of the end of the filtration section that is proximal to the narrow section, configured to limit formation of emulsion.

31. The microfluidic device of any of claims 22 to 30, wherein the pillars are disposed in an array comprising at least one row.

32. The microfluidic device of claim 31, wherein the array comprises 3 rows with a first row including 4 pillars, a second row including 3 pillars, and a third row including 4 pillars .

33. The microfluidic device of any of claims 22 to 32, wherein the impurities include particles, fibers, reagent precipitates, sample matrices, or a combination thereof.

34. The microfluidic device of any of claims 22 to 33, wherein the impurities include particles with a smallest dimension of 10 to 50 microns.

35. The microfluidic device of any of claims 22 to 34, wherein the fluid includes an oil, water, a nuclei acid, or a combination thereof.

36. The microfluidic device of any of claims 22 to 35, wherein the sample compartments each have a volume of 477 to 583 pico liters.

37. The microfluidic device of any of claims 22 to 36, wherein micro fluidic device comprises at least 10,000 sample compartments.

38. The microfluidic device of any of claims 22 to 37, wherein microfluidic device comprises a plurality of terminating chambers in fluidic communication with the loading conduit and the sample compartments.

39. The microfluidic device of any of claims 22 to 38, wherein the terminating chambers are configured to receive overflown or residual volume of the fluid.

40. The microfluidic device of any of claims 22 to 39, further comprising a plurality of siphon conduits, a siphon conduit of the plurality of siphon conduits in fluidic communication with the loading conduit and a sample compartment.

41. The microfluidic device of claim 40, wherein the siphon conduit is configured to enable fluidic communication between the loading conduit and the sample compartments.

42. The microfluidic device of any of claims 22 to 41, further comprising a thin film applied to the microfluidic device, configured to form a cover for the sample compailments.

43. The microfluidic device of claim 42, wherein the thin film applied to the microfluidic device is further configured to form a cover for at least a portion of the loading conduit.

44. The microfluidic device of any of claims 42 and 43, wherein the thin film comprises a gas -permeable material.

45. The microfluidic device of any of claims 22 to 44, wherein a ratio of a depth of the sample compartment to a minimum distance between two adjacent sample compartments is 3.96 to 3.09.

46. The microfluidic device of any of claims 22 to 45, wherein the microfluidic device is configured to process samples for polymerase chain reaction (PCR).

47. The microfluidic device of any of claims 22 to 45, wherein the microfluidic device is configured to process samples for digital PCR (dPCR).

48. A method of loading a liquid comprising a biological sample into the microfluidic device of any of claims 22 to 47, the method comprising: drawing the liquid to flow through the loading conduit into the sample compartments; wherein the liquid in the sample compartments comprises substantially no impurities with a size of more than 10 microns.

49. The method of claim 48, wherein the liquid further comprises reagents for a PCR process.

50. The method of claim 49, wherein the liquid further comprises an oil, an aqueous solution, a nucleic acid, or combinations thereof.

51. A method of processing a biological sample, the method comprising: applying a plurality of pressure pulses to a liquid mixture comprising the biological sample such that the at least some liquid mixture flows through the loading conduit into the sample compartments of the microfluidic device of any of claims 22 to 47 ; and performing PCR amplification by thermal cycling the liquid mixture in the sample compartments.

52. The method of claim 51, wherein the liquid mixture in the sample compartments comprises substantially no impurities with a size over 10 microns.

53. The method of any of claims 51 and 52, further comprising capturing images of the sample compartments of the microfluidic device.

54. The method of claim 53, further comprising determining a number of sample compartments within which PCR amplification has been successfully achieved based on the images of the sample compartments of the microfluidic device.