Devices and systems for automated assays and uses thereof

Abstract

To provide devices and systems for performing automated multiplex assays, methods of using such devices and systems, and a magnetic module for reacting or labeling magnetic microcarriers.SOLUTION: Microcarriers are initially contained in a microcarrier well in a dry form, and suspended by transferring a sample (e.g., amplicon sample) to the microcarrier well and / or by transferring a hybridization buffer from a hybridization buffer well to the microcarrier well using a liquid handling module.SELECTED DRAWING: Figure 1

Description

[Technical field]

[0001] (CROSS REFERENCE TO RELATED APPLICATIONS) This application claims priority to U.S. Provisional Application No. 63 / 514,771, filed July 20, 2023, the contents of which are incorporated herein by reference in their entirety.

[0002] The present disclosure generally relates to devices and systems for performing automated multiplex assays and methods of using the same. In some embodiments, the devices and systems include a magnetic module that simplifies and automates the processing of magnetic microcarriers for molecular and immunoassays. [Background technology]

[0003] Immunologic and molecular diagnostic assays play an important role in both research and clinical fields. It is often necessary to perform assays on a panel of multiple targets to obtain a meaningful or panoramic view of the results and to facilitate research or clinical decisions. This is especially true in the era of genomics and proteomics, where an abundance of genetic markers and / or biomarkers are believed to be able to influence or predict certain disease states. In theory, assays for multiple targets can be achieved by testing each target separately in parallel or sequentially in different reaction vessels (i.e., performing multiple singleplexes). However, assays employing singleplexing strategies are often laborious and usually require large amounts of sample, especially when the targets to be analyzed are numerous.

[0004] A multiplex assay measures multiple (two or more) analytes simultaneously in a single assay. Multiplex assays are typically used in high-throughput screening settings where many samples can be analyzed at once. The ability to assay many analytes simultaneously and in parallel is the hallmark of a multiplex assay and what makes it such a powerful tool in fields ranging from drug discovery to functional genomics to clinical diagnostics. Contrary to singleplexing, mixing all targets in the same reaction vessel makes the assay much less laborious and much easier to perform, since only one reaction vessel per sample is handled. Thus, the amount of test sample required can be dramatically reduced, which is especially important when it is difficult and / or invasive to obtain large amounts of sample (e.g., tumor tissue, cerebrospinal fluid, or bone marrow). Equally important, the cost of reagents can be reduced and the throughput of the assay can be dramatically increased.

[0005] Existing systems for multiplex assays or for processing microcarriers generally require complex and cumbersome structural elements such as aspiration channels, dispensing channels, shaking platforms and / or magnetic plates. The systems are generally difficult to maintain, for example to avoid contamination or clogging (e.g., due to crystallization) in the aspiration and dispensing channels, and may occupy excessive space. Also, the use of a shaking platform may consume more energy and generate more noise compared to more localized mixing methods. Thus, there is a need for small, simple and / or easy-to-maintain assay systems, for example, those that do not require aspiration channels, dispensing channels and / or shaking platforms to process microcarriers. Summary of the Invention

[0006] In one aspect, a magnetic module for reacting or labeling magnetic microcarriers is provided herein. In some embodiments, the module comprises one or more well sets. In some embodiments, each well set comprises a reaction well. In some embodiments, each well set comprises one or more reaction wash wells, including a first reaction wash well and a final reaction wash well, each of which comprises a reaction wash buffer. In some embodiments, if there is only one reaction wash well in the well set, the first reaction wash well and the final reaction wash well are the same well. In some embodiments, each well set comprises a label well comprising a label molecule in a label buffer. In some embodiments, each well set comprises one or more label wash wells, including a first label wash well and a final label wash well, each of which comprises a label wash buffer. In some embodiments, if there is only one label wash well in the well set, the first label wash well and the final label wash well are the same well. In some embodiments, the module comprises one or more magnetic units. In some embodiments, each magnetic unit is configured to move into, out of, and between each of the wells of the well set. In some embodiments, each magnetic unit attracts the magnetic microcarriers and moves them into, out of, and between each of the wells of the well set. In some embodiments, each magnetic unit comprises an upper end and a lower end. In some embodiments, the module comprises one or more magnetic unit pockets. In some embodiments, each magnetic unit pocket is positioned between each magnetic unit and the well set. In some embodiments, each magnetic unit pocket is configured to move into, out of, and between each of the wells of the well set. In some embodiments, each magnetic unit pocket is configured to at least partially cover the lower end of each magnetic unit as the magnetic unit moves or slides into the magnetic unit pocket. In some embodiments, each magnetic unit pocket comprises an inner surface and an outer surface. In some embodiments, the magnetic microcarriers are bound to one or more capture agents.In some embodiments, the one or more capture agents are capable of binding to one or more analytes.

[0007] In some embodiments, each well set further comprises an antibody well comprising a secondary antibody in an antibody buffer. In some embodiments, the secondary antibody is capable of binding to one or more analytes and is bound to one or more secondary binding moieties. In some embodiments, each well set further comprises one or more antibody wash wells comprising a first antibody wash well and a final antibody wash well, each of which comprises an antibody wash buffer. In some embodiments, if there is only one antibody wash well in a well set, the first antibody wash well and the final antibody wash well are the same well.

[0008] In some embodiments, the reaction well, one or more reaction wash wells, antibody well, one or more antibody wash wells, label well and one or more label wash wells are positioned in that order along a straight line or a curve. In some embodiments, the secondary antibody is selected from the group consisting of an antibody and an antibody fragment. In some embodiments, the one or more secondary binding moieties are capable of binding to a label molecule. In some embodiments, the one or more secondary binding moieties comprise biotin.

[0009] In some embodiments, the reaction well, the one or more reaction wash wells, the label well, and the one or more label wash wells are positioned in this order along a straight line or a curve. In some embodiments, each of the one or more capture agents is independently selected from the group consisting of small molecules, polynucleotides, polypeptides, proteins, lipids, and polysaccharides. In some embodiments, each of the one or more analytes is independently selected from the group consisting of small molecules, polynucleotides, polypeptides, proteins, lipids, and polysaccharides.

[0010] In some embodiments, each of the one or more capture agents and the one or more analytes are independently selected from the group consisting of polynucleotides. In some embodiments, each of the one or more capture agents is independently selected from the group consisting of antibodies and antibody fragments, and each of the one or more analytes is independently selected from the group consisting of antigens. In some embodiments, each of the one or more capture agents is independently selected from the group consisting of antigens, and each of the one or more analytes is independently selected from the group consisting of antibodies.

[0011] In some embodiments, the label molecule comprises a label binding moiety attached to a report moiety. In some embodiments, the label binding moiety is streptavidin (SA) or an anti-biotin antibody. In some embodiments, the report moiety is phycoerythrin (PE) or fluorescein isothiocyanate (FITC).

[0012] In some embodiments, each magnetic unit has a rod-like shape. In some embodiments, each magnetic unit pocket has a bottomed cylindrical shape. In some embodiments, one or more magnetic unit pockets are configured to move or vibrate vertically relative to one or more well sets. In some embodiments, one or more magnetic unit pockets are disposable. In some embodiments, the number of one or more magnetic units is the same as the number of one or more magnetic unit pockets. In some embodiments, the number of one or more magnetic units is the same as the number of reaction wells.

[0013] In some embodiments, the various wells are positioned along a line, i.e., form a one-dimensional array. In some embodiments, the reaction wells are positioned along a line, or the label wells are positioned along a line, or both. In some embodiments, one or more well sets form a multiwell plate, with each well set forming a row of the multiwell plate, and the various wells form a column of the multiwell plate. In some embodiments, one or more magnetic units are positioned along a straight line above the multiwell plate and parallel to the column direction of the multiwell plate. In some embodiments, one or more magnetic unit pockets are positioned along a straight line between the multiwell plate and the one or more magnetic units and parallel to the column direction of the multiwell plate. In some embodiments, the various wells are positioned along a surface, i.e., form a two-dimensional array. In some embodiments, the reaction wells are positioned along a surface, i.e., form a two-dimensional array, or the label wells are positioned along a surface, i.e., form a two-dimensional array, or both. In some embodiments, the various wells form a multiwell plate, such that one or more well sets comprise a plurality of multiwell plates. In some embodiments, the one or more magnetic units and the various wells are in the same one or two dimensional array such that one or more magnetic units can move into and out of the various wells at the same time. In some embodiments, the one or more magnetic unit pockets and the various wells are in the same one or two dimensional array such that one or more magnetic unit pockets can move into and out of the various wells at the same time.

[0014] In some embodiments, the reaction well comprises the coded microcarriers or a reaction buffer or both. In some embodiments, the magnetic microcarriers comprise coded microcarriers, spherical magnetic beads or a combination thereof. In some embodiments, each of the coded microcarriers comprises a substantially transparent magnetic polymer layer having a first side and a second side, the first and second sides being parallel to each other, the substantially transparent magnetic polymer comprising a mixture of a substantially transparent polymer and a plurality of magnetic nanoparticles, the magnetic nanoparticles comprising iron (II, III) oxide or iron (III) oxide. In some embodiments, each of the coded microcarriers comprises a substantially opaque layer, the substantially opaque layer being immobilized on the first side of the substantially transparent magnetic polymer layer, and the contours of the substantially opaque layer constitute a two-dimensional shape representing the analog code. In some embodiments, each of the coded microcarriers comprises one or more capture agents for capturing one or more analytes, the one or more capture agents being bound to at least one of the first side and the second side of the substantially transparent magnetic polymer layer. In some embodiments, the magnetic component of the coded microcarriers comprises a substantially transparent magnetic polymer layer. In some embodiments, the magnetic nanoparticles are superparamagnetic. In some embodiments, the magnetic nanoparticles are less than about 30 nm in diameter and greater than about 3 nm in diameter. In some embodiments, the plurality of magnetic nanoparticles comprises less than about 10% (by weight) and greater than about 0.1% (by weight) of a mixture of a substantially transparent polymer and a plurality of magnetic nanoparticles. In some embodiments, the substantially transparent magnetic polymer layer is about 0.1 μm to about 50 μm thick. In some embodiments, the substantially transparent polymer is an epoxy-based polymer. In some embodiments, the epoxy-based polymer is SU-8. In some embodiments, the substantially opaque layer comprises a substantially opaque polymer. In some embodiments, the substantially opaque layer comprises a black matrix resist. In some embodiments, the substantially opaque polymer exhibits an absorptivity greater than about 1.8 (OD) at wavelengths from about 230 nm to about 660 nm. In some embodiments, the substantially opaque layer comprises a metal with no remanent magnetism. In some embodiments, the substantially opaque layer comprises titanium or chromium.In some embodiments, the substantially opaque layer is about 0.05 μm to about 2 μm thick. In some embodiments, the analog code comprises one or more overlapping or partially overlapping arc elements forming a continuous or discontinuous ring. In some embodiments, each of the coded microcarriers comprises an orientation indicator for orienting the analog code of the substantially opaque layer. In some embodiments, the orientation indicator comprises an asymmetry of the substantially opaque layer. In some embodiments, the microcarrier is a generally circular disk. In some embodiments, the microcarrier is about 5 μm to about 200 μm in diameter, or the microcarrier is about 40 μm in diameter. In some embodiments, the microcarrier is less than about 50 μm thick, the microcarrier is about 2 μm to 10 μm thick, or the microcarrier is about 5 μm thick. In some embodiments, each of the one or more analytes and the one or more capture agents is independently selected from the group consisting of DNA molecules, DNA-like molecules, RNA molecules, RNA-like molecules, polynucleotides, proteins, enzymes, lipids, phospholipids, carbohydrate moieties, polysaccharides, antigens, viruses, cells, antibodies, small molecules, bacterial cells, organelles, and antibody fragments.

[0015] In some aspects, an assay system is provided herein that includes a magnetic module as described herein. In some embodiments, the system further includes a detection module configured to detect a signal from the magnetic microcarriers. In some embodiments, the detection module includes one or more detection wells. In some embodiments, the one or more detection wells have a flat bottom. In some embodiments, the one or more detection wells include a final labeling wash well. In some embodiments, the one or more detection wells and the one or more magnetic units have the same one-dimensional or two-dimensional arrangement, such that one or more magnetic units can move into one or more detection wells at the same time and move out of one or more detection wells at the same time. In some embodiments, the one or more detection wells and the one or more magnetic unit pockets have the same one-dimensional or two-dimensional arrangement, such that one or more magnetic unit pockets can move into one or more detection wells at the same time and move out of one or more detection wells at the same time.

[0016] In some embodiments, the assay system further comprises a PCR module configured to perform a polymerase chain reaction (PCR). In some embodiments, the PCR module is configured to perform a PCR using primers bound to one or more secondary binding moieties capable of binding to the target molecule. In some embodiments, the secondary binding moiety comprises biotin.

[0017] In some embodiments, the assay system further comprises a liquid handling module. In some embodiments, the liquid handling module is configured to transfer the sample from the PCR module to the magnetic module. In some embodiments, the liquid handling module is configured to transfer the sample from the PCR module to a reaction well of the magnetic module.

[0018] In some embodiments, each well set further comprises a hybridization buffer well containing a hybridization buffer, and the liquid handling module is configured to transfer at least a portion of the hybridization buffer from the hybridization buffer well to the reaction well. In some embodiments, each well set further comprises an incubation buffer well containing an incubation buffer, and the liquid handling module is configured to transfer at least a portion of the incubation buffer from the incubation buffer well to the reaction well.

[0019] In some aspects, methods of using the magnetic modules or assay systems described herein are provided herein. In some embodiments, the methods comprise (a) transferring one or more analytes into a reaction well, the reaction well containing magnetic microcarriers in a reaction buffer. In some embodiments, the methods comprise (b) moving one of the one or more magnetic unit pockets into the reaction well and moving or vibrating the magnetic unit pocket within the reaction well to mix the one or more analytes with the magnetic microcarriers. In some embodiments, the methods comprise (c) moving the magnetic unit into the reaction well and into the magnetic unit pocket to attract the magnetic microcarriers to an outer surface of the magnetic unit pocket. In some embodiments, the methods comprise (d) transferring the magnetic microcarriers from the reaction well to the first reaction wash well by moving the magnetic unit pocket and the magnetic unit out of the reaction well and into the first reaction wash well. In some embodiments, the methods comprise (e) transferring the magnetic microcarriers from the final reaction wash well to the label well by moving the magnetic unit pocket and the magnetic unit out of the final reaction wash well and into the label well. In some embodiments, the method comprises (f) moving the magnetic unit out of the labeling well while retaining the magnetic unit pocket within the labeling well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the labeling buffer. In some embodiments, the method comprises (g) moving the magnetic unit into the labeling well and into the magnetic unit pocket to attract the magnetic microcarriers to the outer surface of the magnetic unit pocket. In some embodiments, the method comprises (h) transferring the magnetic microcarriers from the labeling well to the first labeling washing well by moving the magnetic unit pocket and magnetic unit out of the labeling well and into the first labeling washing well. In some embodiments, steps (a) through (h) are performed in the order listed.In some embodiments, the method further comprises, after step (d) and before step (e), moving the magnetic unit out of the first reaction wash well while holding the magnetic unit pocket in the first reaction wash well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the first reaction wash well. In some embodiments, the method further comprises, after step (d) and before step (e), moving or vibrating the magnetic unit pocket in the first reaction wash well to mix the magnetic microcarriers with the reaction wash buffer. In some embodiments, the method further comprises, after step (f) and before step (g), moving or vibrating the magnetic unit pocket in the label well to mix the magnetic microcarriers with the label buffer. In some embodiments, the method further comprises, after step (h), moving the magnetic unit out of the first label wash well while holding the magnetic unit pocket in the first label wash well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the first label wash well. In some embodiments, the method further comprises, after step (h), moving or vibrating the magnetic unit pocket in the first label wash well to mix the magnetic microcarriers with the label wash buffer. In some embodiments, the method further comprises, after step (h), transferring the magnetic microcarriers from the final reaction wash well to the detection well by moving the magnetic unit pocket and magnetic unit out of the final label wash well and into the detection well. In some embodiments, the method further comprises moving the magnetic unit out of the detection well while retaining the magnetic unit pocket in the detection well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the detection well. In some embodiments, the method further comprises, after step (h), optically reading the magnetic microcarriers and detecting a signal from the label molecule bound to the magnetic microcarriers.

[0020] In some embodiments, the method comprises (a) transferring one or more analytes into a reaction well, the reaction well containing magnetic microcarriers in a reaction buffer. In some embodiments, the method comprises (b) moving one of the one or more magnetic unit pockets into the reaction well and moving or vibrating the magnetic unit pocket within the reaction well to mix the one or more analytes with the magnetic microcarriers. In some embodiments, the method comprises (c) moving the magnetic unit into the reaction well and into the magnetic unit pocket to attract the magnetic microcarriers to an outer surface of the magnetic unit pocket. In some embodiments, the method comprises (d) transferring the magnetic microcarriers from the reaction well to the first reaction wash well by moving the magnetic unit pocket and magnetic unit out of the reaction well and into the first reaction wash well. In some embodiments, the method comprises (e) transferring the magnetic microcarriers from the final reaction wash well to the antibody well by moving the magnetic unit pocket and magnetic unit out of the final reaction wash well and into the antibody well. In some embodiments, the method comprises (f) moving the magnetic unit out of the antibody well while retaining the magnetic unit pocket within the label well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the antibody buffer. In some embodiments, the method comprises (g) moving the magnetic unit into the antibody well and into the magnetic unit pocket to attract the magnetic microcarriers to the outer surface of the magnetic unit pocket. In some embodiments, the method comprises (h) transferring the magnetic microcarriers from the antibody well to the first antibody washing well by moving the magnetic unit pocket and magnetic unit out of the antibody well and into the first antibody washing well. In some embodiments, the method comprises (i) transferring the magnetic microcarriers from the final antibody washing well to the label well by moving the magnetic unit pocket and magnetic unit out of the final antibody washing well and into the label well.In some embodiments, the method comprises (j) moving the magnetic unit out of the labeling well while retaining the magnetic unit pocket within the labeling well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the labeling buffer. In some embodiments, the method comprises (k) moving the magnetic unit into the labeling well and into the magnetic unit pocket to attract the magnetic microcarriers to the outer surface of the magnetic unit pocket. In some embodiments, the method comprises (l) transferring the magnetic microcarriers from the labeling well to the first labeling wash well by moving the magnetic unit pocket and the magnetic unit out of the labeling well and into the first labeling wash well. In some embodiments, steps (a) through (l) are performed in the order listed. In some embodiments, the method comprises, after step (d) and before step (e), moving the magnetic unit out of the first reaction wash well while retaining the magnetic unit pocket within the first reaction wash well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the first reaction wash well. In some embodiments, the method comprises, after step (d) and before step (e), moving or vibrating the magnetic unit pocket in the first reaction wash well to mix the magnetic microcarriers with the reaction wash buffer. In some embodiments, the method comprises, after step (f) and before step (g), moving or vibrating the magnetic unit pocket in the antibody well to mix the magnetic microcarriers with the labeling buffer. In some embodiments, the method comprises, after step (h) and before step (i), moving the magnetic unit out of the first antibody wash well while retaining the magnetic unit pocket in the first antibody wash well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the first antibody wash well. In some embodiments, the method comprises, after step (h) and before step (i), moving or vibrating the magnetic unit pocket in the first antibody wash well to mix the magnetic microcarriers with the antibody wash buffer.In some embodiments, the method comprises, after step (j) and before step (k), moving or vibrating the magnetic unit pocket in the label well to mix the magnetic microcarriers with the label buffer. In some embodiments, the method comprises, after step (l), moving the magnetic unit out of the first label wash well while retaining the magnetic unit pocket in the first label wash well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the first label wash well. In some embodiments, the method comprises, after step (l), moving or vibrating the magnetic unit pocket in the first label wash well to mix the magnetic microcarriers with the label wash buffer. In some embodiments, the method comprises, after step (l), transferring the magnetic microcarriers from the final reaction wash well to the detection well by moving the magnetic unit pocket and the magnetic unit out of the final label wash well and into the detection well. In some embodiments, the method comprises moving the magnetic unit out of the detection well while retaining the magnetic unit pocket in the detection well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the detection well. In some embodiments the method comprises after step (l) optically reading the magnetic microcarriers to detect signals from labelled molecules bound to the magnetic microcarriers. [Brief description of the drawings]

[0021] [Figure 1] FIG. 1 illustrates an exemplary magnetic module. [Diagram 2] FIG. 2 shows an exemplary assay system. [Diagram 3] FIG. 3 shows an exemplary set of wells where the various wells form rows of a multiwell plate. [Figure 4A] FIG. 4A shows an exemplary set of wells where the various wells form a multi-well plate. [Figure 4B] FIG. 4B shows an exemplary set of wells where the various wells form a multi-well plate. [Figure 4C]FIG. 4C shows an exemplary set of wells where the various wells form a multi-well plate. [Figure 4D] FIG. 4D shows an exemplary set of wells where the various wells form a multi-well plate. [Figure 4E] FIG. 4E shows an exemplary set of wells where the various wells form a multi-well plate. [Figure 4F] FIG. 4F shows an exemplary set of wells where the various wells form a multi-well plate. [Diagram 5] FIG. 5 shows an exemplary process for a molecular assay using the magnetic modules provided herein. [Figure 6] FIG. 6 shows an exemplary design of a well set for a molecular assay using the magnetic modules provided herein. [Figure 7] FIG. 7 shows an exemplary process for an antigen immunoassay using the magnetic modules provided herein. [Figure 8] FIG. 8 shows an exemplary design of a well set for an immunoassay (for either antigen or antibody detection) using the magnetic modules provided herein. [Figure 9] FIG. 9 shows an exemplary process for an antibody immunoassay using the magnetic modules provided herein. [Figure 10] FIG. 10 shows the results of an antigen immunoassay performed by the magnetic module provided herein. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0022] The present application can be understood by reference to the following description taken in conjunction with the accompanying drawings.

[0023] The following description is presented to enable those skilled in the art to make and use the various embodiments. Descriptions of specific devices, techniques, and applications are provided only as examples. Various modifications to the examples described herein will be readily apparent to those skilled in the art, and the general principles defined herein may be applied to other examples and applications without departing from the spirit and scope of the various embodiments. Thus, the various embodiments are not intended to be limited to the examples described and shown herein, but are intended to be accorded the scope consistent with the appended claims.

[0024] I. Definition Before describing the invention in detail, it is to be understood that this invention is not limited to particular compositions or biological systems, which can, of course, vary. Also, it is to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting.

[0025] As used herein, the term "microcarrier" may refer to a physical substrate to which a capture agent may be attached. Microcarriers of the present disclosure may take any suitable geometric form or shape. In some embodiments, the microcarrier may be in the shape of a disk. Typically, the form or shape of the microcarrier is within the range of 10 -4 ~10 -7 It will have at least one dimension on the order of m (hence the prefix "micro").

[0026] As used herein, the term "polymer" may refer to any macromolecular structure that includes repeating monomers. A polymer may be natural (e.g., found in nature) or synthetic (e.g., man-made, such as a polymer made of non-natural monomers and / or polymerized in configurations or combinations not found in nature). As used herein, the terms "substantially transparent" and "substantially opaque" may refer to the ability of light (e.g., of a particular wavelength, such as infrared, visible, UV, etc.) to pass through a substrate, such as a polymer layer. A substantially transparent polymer may refer to one that is transparent, translucent, and / or transmissive to light, while a substantially opaque polymer may refer to one that reflects and / or absorbs light. It should be understood that whether a material is substantially transparent or substantially opaque may depend on the wavelength and / or intensity of the light irradiating the material and the manner in which the light (or its reduction or absence) traveling through the material is detected. In some embodiments, a substantially opaque material causes a perceptible reduction in transmitted light compared to surrounding materials or image areas, for example, as imaged by optical microscopy (e.g., bright field, dark field, phase contrast, differential interference contrast (DIC), Nomarski interference microscopy (NIC), Nomarski microscopy, Hoffman modulation contrast microscopy (HMC), or fluorescence microscopy). In some embodiments, a substantially transparent material allows a perceptible amount of transmitted light to pass through the material, for example, as imaged by optical microscopy (e.g., bright field, dark field, phase contrast, differential interference contrast (DIC), Nomarski interference microscopy (NIC), Nomarski microscopy, Hoffman modulation contrast microscopy (HMC), or fluorescence microscopy).

[0027] As used herein, the term "analog code" may refer to any code in which the coded information is represented in a non-quantized and / or non-discrete manner, e.g., in contrast to digital codes. For example, digital codes are sampled at discrete locations for a limited set of values ​​(e.g., 0 / 1 type values), whereas analog codes may be sampled at a wider range of locations (or as a continuous whole) and / or may include a wider set of values ​​(e.g., shapes). In some embodiments, analog codes may be read or decoded using one or more analog shape recognition techniques.

[0028] As used herein, the term "capture agent" is a broad term and is used in its ordinary sense to refer to any compound or substance capable of specifically recognizing an analyte of interest. In some embodiments, specific recognition may refer to specific binding. Non-limiting examples of capture agents include, for example, DNA molecules, DNA-like molecules, RNA molecules, RNA-like molecules, polynucleotides, proteins, enzymes, lipids, phospholipids, carbohydrate moieties, polysaccharides, antigens, viruses, cells, antibodies, small molecules, bacterial cells, organelles, and antibody fragments.

[0029] As used herein, "analyte" is a broad term and is used in its ordinary sense as a substance whose presence, absence, or amount is to be determined, and is taken to refer, without limitation, to a substance or chemical constituent in a sample, such as an analyzable biological sample or a cell or population of cells. An analyte can be a substance for which a naturally occurring binding member exists or a substance for which a binding member can be prepared. Non-limiting examples of analytes include, for example, antibodies, antibody fragments, antigens, polynucleotides (such as DNA molecules, DNA-like molecules, RNA molecules, or RNA-like molecules), polypeptides, proteins, enzymes, lipids, phospholipids, carbohydrate moieties, polysaccharides, small molecules, organelles, hormones, cytokines, growth factors, steroids, vitamins, toxins, drugs, and metabolites of the above substances, as well as cells, bacteria, viruses, fungi, algae, fungal spores, and the like.

[0030] The term "antibody" is used in the broadest sense and includes monoclonal antibodies (including full-length antibodies having an immunoglobulin Fc region), polyclonal antibodies, multispecific antibodies (e.g., bispecific antibodies, diabodies and single chain molecules) and antibody fragments (e.g., Fab, F(ab')2 and Fv).

[0031] As used herein, a "sample" refers to a composition that contains material, such as a molecule, to be detected. In one embodiment, a sample is a "biological sample" (i.e., any material obtained from a living source (e.g., human, animal, plant, bacteria, fungus, protist, virus)). Biological samples can be in any form, including solid materials (e.g., tissues, cell pellets, and biopsies) and biological fluids (e.g., urine, blood, saliva, lymph, tears, sweat, prostatic fluid, seminal fluid, semen, bile, mucus, amniotic fluid, and mouthwash (containing buccal cells)). Solid materials are usually mixed with a liquid. A sample may also refer to an environmental sample, such as water, air, soil, or any other environmental source.

[0032] As used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural references unless the context clearly dictates otherwise. Thus, for example, a reference to "a molecule" optionally includes a combination of two or more such molecules, and so forth.

[0033] As used herein, the term "about" refers to a normal error range for the respective value that is readily apparent to one of ordinary skill in the art. Reference herein to a value or parameter with "about" includes (and describes) an embodiment directed to that value or parameter itself.

[0034] It is understood that aspects and embodiments of the invention described herein include "comprising," "consisting of," and "consisting essentially of" aspects and embodiments.

[0035] II. Magnetic Module In one aspect, a magnetic module is described herein. In some embodiments, the magnetic module is for reacting or labeling magnetic microcarriers. In some embodiments, the magnetic module comprises one or more well sets, one or more magnetic units, one or more magnetic unit pockets, or any combination thereof. In some embodiments, the magnetic microcarriers comprise a magnetic component. In some embodiments, the magnetic microcarriers are bound to one or more capture agents, and the one or more capture agents can bind to one or more analytes.

[0036] In some embodiments, one or more of the well sets comprise a reaction well. In some embodiments, the reaction well is configured to comprise a reaction buffer. Any suitable reaction buffer can be used. In some embodiments, the reaction buffer is a hybridization buffer. In some embodiments, the reaction buffer is an incubation buffer. In some embodiments, the reaction well is configured to comprise one or more magnetic microcarriers. In some embodiments, the reaction well is an empty well. In some embodiments, the reaction well is configured to comprise both a reaction buffer and a magnetic microcarrier. In some embodiments, the magnetic microcarrier comprises an encoded microcarrier as described herein. In some embodiments, the magnetic microcarrier comprises a magnetic latex bead. In some embodiments, the magnetic microcarrier comprises a spherical magnetic bead. In some embodiments, the magnetic microcarrier comprises a spherical magnetic latex bead.

[0037] In some embodiments, the one or more well sets comprise one or more reaction wash wells. In some embodiments, the one or more reaction wash wells comprise a first reaction wash well and a final reaction wash well, each of which is configured to contain a reaction wash buffer. In some embodiments, there is only one reaction wash well in a well set, and the first reaction wash well and the final reaction wash well are the same well.

[0038] In some embodiments, one or more of the well sets comprises a label well. In some embodiments, the label well contains a label molecule in a labeling buffer. In some embodiments, the label molecule is a streptavidin-phycoerythrin (PE) conjugate.

[0039] In some embodiments, the one or more well sets comprise one or more label wash wells. In some embodiments, the one or more label wash wells comprise a first label wash well and a final label wash well, each of which is configured to contain a label wash buffer. In some embodiments, there is only one label wash well in a well set, and the first label wash well and the final label wash well are the same well.

[0040] In some variations, one or more of the well sets further comprise wells that contain additional reagents, such as an antibody well and one or more antibody wash wells.

[0041] In some embodiments, the various wells in one or more well sets form a one-dimensional array. The one-dimensional array can be a straight line, a set of straight lines connected at their ends to each other, or a curve. FIG. 3 shows an exemplary 8×9 array of wells. A set of wells (well sets) comprising one of the various wells (i.e., reaction well, first reaction wash well, second reaction wash well, final reaction wash well, label well, first label wash well, second label wash well, third label wash well, and final label wash well) forms a row of a matrix array. In some embodiments, the reaction wells are positioned along a line (e.g., FIG. 3). In some embodiments, the label wells are positioned along a line (e.g., FIG. 3). In some embodiments, the one or more well sets form a multiwell plate (e.g., FIG. 3). In Figure 3, each type of well (i.e., reaction well, first reaction wash well, second reaction wash well, final reaction wash well, label well, first label wash well, second label wash well, third label wash well, or final label wash well) forms each column of the 8x9 multiwell plate, and each row comprising one of each type of well forms a well set. In some embodiments, each well set forms each row of the multiwell plate (e.g., Figure 3), and each type of well forms each column of the multiwell plate (e.g., Figure 3).

[0042] In some embodiments, the various wells form a two-dimensional array. The two-dimensional array can be along a surface (such as a multiwell plate). In some embodiments, the two-dimensional array is a two-dimensional array. The array can be any shape, such as a square, rectangle, triangle, circle, or hexagon. In some embodiments, the various wells form a multiwell plate. In some embodiments, one or more well sets comprise a plurality of multiwell plates. FIGS. 4A-4F show exemplary well sets comprising a two-dimensional array in the form of a multiwell plate. FIG. 4A shows a multiwell plate with reaction wells positioned along a flat surface to form an 8×12 two-dimensional array. FIG. 4B shows a multiwell plate with a first reaction wash well positioned along a flat surface to form an 8×12 two-dimensional array. FIG. 4C shows a multiwell plate with a last reaction wash well positioned along a flat surface to form an 8×12 two-dimensional array. FIG. 4D shows a multiwell plate with label wells positioned along a flat surface to form an 8×12 two-dimensional array. Figure 4E shows a multiwell plate with the first labeled wash wells positioned along the flat surface to form an 8 x 12 two-dimensional array. Figure 4F shows a multiwell plate with the last labeled wash wells positioned along the flat surface to form an 8 x 12 two-dimensional array.

[0043] In some embodiments, the magnetic module comprises one or more magnetic units having an upper end and a lower end. In some embodiments, each magnetic unit is configured to move into, out of, and between each of the wells of the well set. In some embodiments, the magnetic module comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, In some embodiments, the magnetic module comprises 0, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96 or more magnetic units. In some embodiments, the magnetic module comprises 6, 12, 24, 48, 96, 192 or 384 magnetic units. In some embodiments, the magnetic module comprises 8 magnetic units. In some embodiments, the magnetic module comprises 96 magnetic units. In some embodiments, the magnetic module is configured to attract magnetic microcarriers.

[0044] In some embodiments, the magnetic module comprises one or more magnetic unit pockets comprising an inner surface and an outer surface. The magnetic module may comprise 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96 or more magnetic unit pockets. In some embodiments, the magnetic module comprises 6, 12, 24, 48, 96, 192 or 384 magnetic unit pockets. In some embodiments, the magnetic module comprises 8 magnetic unit pockets. In some embodiments, the magnetic module comprises 96 magnetic unit pockets.

[0045] In some embodiments, one or more magnetic units, one or more magnetic unit pockets, and various wells are in the same one-dimensional or two-dimensional array. For example, the magnetic units, magnetic unit pockets, and label wells are in an 8×1 array (i.e., 8 magnetic units make up a line, 8 magnetic unit pockets form a line, and 8 label wells form a line). In some embodiments, the magnetic units, magnetic unit pockets, and label wells are in an 8×12 array (i.e., 96 magnetic units form an 8×12 array, 96 magnetic unit pockets form an 8×12 array, and 96 label wells form an 8×12 array). In some embodiments, one or more magnetic units and one or more magnetic unit pockets are in the same one-dimensional or two-dimensional array. In some embodiments, one or more magnetic units and various wells are in the same one-dimensional or two-dimensional array. In some embodiments, one or more magnetic unit pockets and various wells are in the same one-dimensional or two-dimensional array.

[0046] In some embodiments, the magnetic units and magnetic unit pockets can move into the various wells at the same time and out of the various wells at the same time. In some embodiments, the magnetic units can move into the various wells at the same time and out of the various wells at the same time. In some embodiments, the magnetic unit pockets can move into the various wells at the same time and out of the various wells at the same time. In some embodiments, the magnetic units can move into the various magnetic unit pockets at the same time and out of the various magnetic unit pockets at the same time.

[0047] In some embodiments, the magnetic unit pocket is positioned between each magnetic unit and the set of wells. In some embodiments, the magnetic unit pocket is configured to at least partially cover the bottom end of each magnetic unit as the magnetic unit moves or slides into the magnetic unit pocket. In some embodiments, the number of the one or more magnetic units is the same as the number of the one or more magnetic unit pockets. In some embodiments, the number of the one or more magnetic units is the same as the number of wells of each type. In some embodiments, the number of the one or more magnetic units is the same as the number of reaction wells. In some embodiments, the number of the one or more magnetic units is the same as the number of label wells. In some embodiments, the one or more magnetic units are positioned along a straight line above the multiwell plate. In some embodiments, the one or more magnetic units are positioned parallel to the column direction of the multiwell plate, such as the multiwell plate shown in FIG. 3.

[0048] FIG. 1 shows an exemplary magnetic module 100 comprising a magnetic unit gantry 102, a magnetic unit 104, a pocket gantry 106, a magnetic unit pocket 108, a position control unit 110 and a well region 112. The magnetic units are mounted to the magnetic unit gantry whose vertical and horizontal positions are controlled by the position control unit. The magnetic unit pocket is connected to the pocket gantry whose vertical and horizontal positions are controlled by the position control unit. The position control unit independently controls the positions of the magnetic unit gantry and the pocket gantry. The magnetic units are positioned above the magnetic unit pocket and configured to move into and out of the magnetic unit pocket. The magnetic unit pocket is configured to at least partially cover the bottom ends of the magnetic units as they move into the magnetic unit pocket. The well region comprises wells in a two-dimensional array.

[0049] In some embodiments, each magnetic unit has a rod-like shape. In some embodiments, each magnetic unit pocket has a bottomed cylindrical shape. In some embodiments, the magnetic unit pocket is configured to move or vibrate vertically relative to the one or more well sets to promote mixing of the buffer and the microcarriers in the wells. In some embodiments, the one or more magnetic unit pockets are reusable. In some embodiments, the one or more magnetic unit pockets are disposable. In some embodiments, the one or more magnetic unit pockets are made of plastic.

[0050] In some aspects, the microcarriers of the present disclosure include a capture agent. In some embodiments, a capture agent for a particular microcarrier type may be a "unique capture agent", e.g., a capture agent is associated with a particular microcarrier type with a specific identifier (e.g., an analog code). A capture agent can be any biomolecule or chemical compound capable of binding one or more analytes (such as biomolecules or chemical compounds) present in a solution. Examples of biomolecular capture agents include, but are not limited to, DNA molecules, DNA-like molecules, RNA molecules, RNA-like molecules, oligonucleotides, polynucleotides, proteins, enzymes, lipids, phospholipids, carbohydrate moieties, polysaccharides, antigens, viruses, cells, antibodies, small molecules, bacterial cells, organelles, and antibody fragments. Examples of chemical compound capture agents include, but are not limited to, individual components of a chemical library, small molecules, or environmental toxins (e.g., pesticides or heavy metals). In some embodiments, the capture agent includes a nucleotide. In some embodiments, the capture agent includes an antigen. In some embodiments, the capture agent includes an antibody.

[0051] In some embodiments, the capture agent is bound to the surface of the microcarrier (in some embodiments, at least at the center of the microcarrier surface). In some embodiments, the capture agent can be chemically attached to the microcarrier. In other embodiments, the capture agent can be physically adsorbed to the surface of the microcarrier. In some embodiments, the attachment bond between the capture agent and the microcarrier surface can be a covalent bond. In other embodiments, the attachment bond between the capture agent and the microcarrier surface can be a non-covalent bond, including but not limited to a salt bridge or other ionic bond, one or more hydrogen bonds, hydrophobic interactions, van der Waals forces, London dispersion forces, mechanical bonds, one or more halogen bonds, lipophilicity, intercalation, or stacking.

[0052] In some aspects, two or more (e.g., 2, 3, 4, 5, 6, 7, 8, 9, or 10) capture agents for the same analyte may each be associated with a microcarrier as described herein. In this embodiment, each capture agent for a particular analyte binds to the analyte with a different affinity as measured by the dissociation constant of the analyte / capture agent binding. Thus, within a plurality of microcarriers in a composition, there may be two or more subpopulations of microcarriers having capture agents that bind to the same analyte, but the capture agents associated with each subpopulation bind to the analyte with a different affinity. In some embodiments, the dissociation constant of the analyte for any of the capture agents is greater than or equal to 10 -7 M~10 -8 M, etc., 10 -6 In other embodiments, the dissociation constant of the analyte with any of the capture agents is about 10 -10 M ~ about 10 -7 M, about 10 -10 M ~ about 10 -8 M, about 10 -10 M ~ about 10 -9 M, about 10 -9 M ~ about 10 -6 M, about 10 -9 M ~ about 10 -7 M, about 10 -9 M ~ about 10 -8 M, about 10 -8 M ~ about 10 -6 M, or about 10 -8 M ~ about 10 -7 M, etc., about 10 -10 M ~ about 10 -6 M. In some embodiments, the dissociation constant of the analyte with any two capture agents is about 2.5 log 10 , 2log 10 , 1.5log 10 Or 1 log 10 Size of the object, etc., about 3 logs 10 are also different.

[0053] In some embodiments, analytes of the present disclosure are bound to microcarriers for capture of one or more analytes. In some embodiments, one or more analytes may be captured from a sample, such as a biological sample, as described herein. In some embodiments, analytes may include, without limitation, DNA molecules, DNA-like molecules, RNA molecules, RNA-like molecules, polynucleotides, proteins, enzymes, lipids, phospholipids, carbohydrate moieties, polysaccharides, antigens, viruses, cells, antibodies, small molecules, bacterial cells, organelles, and antibody fragments. In other embodiments, the analytes are chemical compounds (such as small molecule chemical compounds) that can bind to a capture agent, such as individual components of a chemical library, small molecules, or environmental toxins.

[0054] In some embodiments, each of the one or more capture agents and the one or more analytes are independently selected from the group consisting of polynucleotides. In some embodiments, each of the one or more capture agents is independently selected from the group consisting of antibodies and antibody fragments, and each of the one or more analytes is independently selected from the group consisting of antigens. In some embodiments, each of the one or more capture agents is independently selected from the group consisting of antigens, and each of the one or more analytes is independently selected from the group consisting of antibodies.

[0055] In some aspects, an analyte in a sample (such as a biological sample) can be labeled with a labeling molecule that can emit a detectable signal upon binding to a capture agent. In some embodiments, the labeling molecule can be colorimetric. In other embodiments, the labeling molecule can be fluorescent, including but not limited to phycoerythrin, blue fluorescent protein, green fluorescent protein, yellow fluorescent protein, cyan fluorescent protein, and derivatives thereof. In other embodiments, the labeling molecule can be fluorescent, including but not limited to phycoerythrin, blue fluorescent protein, green fluorescent protein, yellow fluorescent protein, cyan fluorescent protein, and derivatives thereof. 32 P, 33 P, 22 Na, 36 Cl, 2 H, 3 H, 35 S and 123The labeling molecule may be a radioisotope system, including molecules labeled with I. In other embodiments, the labeling molecule is light-based, including, but not limited to, luciferase (e.g., chemiluminescence systems), horseradish peroxidase, alkaline phosphatase, and derivatives thereof. In some embodiments, biomolecules or chemical compounds present in the sample can be labeled with the labeling molecule prior to contact with the microcarriers. In other embodiments, biomolecules or chemical compounds present in the sample can be labeled with the labeling molecule following contact with the microcarriers. In some embodiments, the labeling molecule comprises a label binding moiety attached to a report moiety. In some embodiments, the label binding moiety is streptavidin (SA). In some embodiments, the label binding moiety is an anti-biotin antibody. In some embodiments, the report moiety is phycoerythrin (PE). In some embodiments, the report moiety is fluorescein isothiocyanate (FITC).

[0056] Modules for molecular assays In some embodiments, the magnetic modules provided herein are configured to perform molecular assays.

[0057] FIG. 2 shows an exemplary assay system 200 comprising a PCR module 210 and a magnetic module 230. The magnetic module 230 comprises three magnetic unit gantries 232, three pocket gantries 234, and a well area 236. The well area comprises six rectangular subareas, each of which can accommodate a multi-well plate pre-filled with buffer. In some embodiments, the cross-section or front view of the gantry has an inverted C-shape to form a C-shaped rail. In some embodiments, the top of the magnetic unit is T-shaped. In some embodiments, the top of the magnetic unit pocket is T-shaped. In some embodiments, the T-shaped magnetic unit or magnetic unit pocket is insertable into a corresponding C-shaped rail. In some embodiments, the gantry comprises a C-shaped rail. In some embodiments, a disposable T-shaped magnetic unit pocket can be easily inserted into or moved out of the C-shaped rail of the pocket gantry.

[0058] FIG. 5 shows an exemplary molecular assay process 500. A biotin-labeled amplicon-containing sample (biotin-labeled target X; the amplicon can be a DNA molecule amplified through PCR using a biotin-labeled primer) is added to a reaction well containing a microcarrier bound to a capture agent with probe X (FIG. 5 "encoded microcarrier"). A magnetic unit pocket moves, moves up and down and / or oscillates in the reaction well to facilitate mixing of the amplicon with the microcarrier and hybridization of target X and probe X (FIG. 5 "hybridization"). The magnetic unit moves into the reaction well and into the magnetic unit pocket to attract the microcarrier to the outer surface of the magnetic unit pocket. The magnetic unit and magnetic unit pocket are transferred to a reaction wash well for washing (FIG. 5, first "wash"). The magnetic unit moves out of the reaction well to release the microcarrier into a wash buffer. The magnetic unit pocket moves, moves up and down and / or oscillates to facilitate mixing in the wash well. The magnetic unit moves back into the magnetic unit pocket and attracts the microcarriers to the outer surface of the magnetic unit pocket. The magnetic unit and magnetic unit pocket move to the labeling well and transfer the microcarriers to the labeling well. The magnetic unit moves out of the labeling well and releases the microcarriers into the labeling buffer, and the magnetic unit pocket moves, bobs and / or vibrates to promote mixing of the microcarriers with the labeling molecule (streptavidin-phycoerythrin; SA-PE) in the well (Figure 5, "SA-PE labeling"). The magnetic unit moves into the labeling well and into the magnetic unit pocket and attracts the microcarriers to the outer surface of the magnetic unit pocket. The magnetic unit and magnetic unit pocket with the attracted microcarriers are transferred to the labeling wash well for washing (Figure 5, second "wash"). After washing, the microcarriers are transferred to the detection well for optical reading of the coded microcarriers and detection of the fluorescent signal from the PE bound to the microcarriers. In some variations, the washing step is repeated with multiple washing wells.

[0059] 6 shows a cross-sectional view of an exemplary well set 600 comprising a microcarrier well 602, a hybridization buffer well 604, a first reaction wash well 606, a second reaction wash well 608, a final reaction wash well 610, a label well 612, a first label wash well 614, a second label wash well 616, a third label wash well 618, and a final label wash well 620. A cross-sectional view of an exemplary T-shaped magnetic unit pocket 622 is also shown. In some embodiments, the coded microcarriers are initially contained in the microcarrier wells in dry form and are suspended by using a liquid handling module to transfer a sample (e.g., an amplicon sample) to the microcarrier well and / or to transfer hybridization buffer from the hybridization buffer well to the microcarrier well. In this case, the microcarrier wells may be reaction wells.

[0060] Modules for immunoassays In some embodiments, the magnetic modules provided herein are configured to perform an immunoassay. In some embodiments, the well set in the magnetic module further comprises an antibody well. In some embodiments, the antibody well comprises a secondary antibody in an antibody buffer. In some embodiments, the secondary antibody is bound to one or more secondary binding moieties. In some embodiments, the secondary antibody is capable of binding to one or more analytes. In some embodiments, each of the well sets in the magnetic module further comprises one or more antibody wash wells, each comprising a first antibody wash well and a final antibody wash well, each comprising an antibody wash buffer. In some embodiments, if there is only one antibody wash well in the set, the first antibody wash well and the final antibody wash well are the same well.

[0061] The wells in a well set may be arranged in any suitable order. In some embodiments, the reaction well, one or more reaction wash wells, antibody well, one or more antibody wash wells, label well and one or more label wash wells are positioned in this order along a straight line. In some embodiments, the reaction well, one or more reaction wash wells, antibody well, one or more antibody wash wells, label well and one or more label wash wells are positioned in this order along a curve. In some embodiments, the number of one or more magnetic units and the number of antibody wells are the same.

[0062] In some embodiments, the secondary antibody is selected from the group consisting of an antibody and an antibody fragment. In some embodiments, the one or more secondary binding moieties are capable of binding to a labeled molecule. In some embodiments, the one or more secondary binding moieties comprise biotin.

[0063] FIG. 7 shows an exemplary immunoassay process 700 for detecting an antigen in a sample. A sample containing an antigen is added to a reaction well containing microcarriers bound to a capture agent that contains an antibody capable of binding to a target antigen (FIG. 7, "Encoded Microcarriers"). The magnetic unit pocket moves into the reaction well, moves up and down, and / or vibrates to promote mixing of the antigen and the microcarriers (FIG. 7, "Incubation"). The magnetic unit moves into the reaction well and into the magnetic unit pocket to attract the microcarriers to the outer surface of the magnetic unit pocket. The magnetic unit and magnetic unit pocket are transferred to a reaction wash well for washing (FIG. 7, first "Wash"). The magnetic unit moves out of the reaction well to release the microcarriers into a wash buffer. The magnetic unit pocket moves up and down, and / or vibrates to promote mixing in the well. The magnetic unit moves back into the magnetic unit pocket to attract the microcarriers to the outer surface of the magnetic unit pocket. The magnetic unit and magnetic unit pocket move to the antibody well. The magnetic unit moves out of the antibody well to release the microcarriers into the antibody buffer, and the magnetic unit pocket moves, moves up and down, and / or vibrates to facilitate mixing of the microcarriers with the secondary antibody in the well (Figure 7, "Biotinylated Detection Ab Incubation"). The secondary antibodies bind to the antigens bound to the microcarriers through their capture agents. The magnetic unit moves into the antibody well and into the magnetic unit pocket to attract the microcarriers to the outer surface of the magnetic unit pocket. The magnetic unit and magnetic unit pocket are transferred to the antibody washing well for washing (Figure 7, second "wash"). The magnetic unit moves out of the antibody washing well to release the microcarriers into the washing buffer. The magnetic unit pocket moves, moves up and down, and / or vibrates to facilitate mixing in the washing well. The magnetic unit moves back into the magnetic unit pocket to attract the microcarriers to the outer surface of the magnetic unit pocket. The magnetic unit and magnetic unit pocket move to the labeling well to transfer the microcarriers to the labeling well.The magnetic unit moves out of the labeling well to release the microcarriers into the labeling buffer, and the magnetic unit pocket moves, bobs and / or vibrates to promote mixing of the microcarriers with the labeling molecule (streptavidin-phycoerythrin; SA-PE) in the well (Figure 7, "SA-PE labeling"). The magnetic unit moves into the labeling well and into the magnetic unit pocket to attract the microcarriers to the outer surface of the magnetic unit pocket. The magnetic unit and magnetic unit pocket, along with the attracted microcarriers, are transferred to the labeling washing well for washing (Figure 7, third "wash"). After washing, the microcarriers are transferred to the detection well for optical reading of the coded microcarriers and detection of the fluorescent signal from the labeling molecule bound to the microcarriers. In some variations, the washing step is repeated in multiple washing wells.

[0064] 8 shows a cross-sectional view of an exemplary well set 800 comprising a microcarrier well 802, an incubation buffer well 804, a reaction wash well 806, an antibody well 808, a first antibody wash well 810, a final antibody wash well 812, a label well 814, a first label wash well 816, a second label wash well 818 and a final label wash well 820. A cross-sectional view of an exemplary T-shaped magnetic unit pocket 822 is also shown. In some embodiments, the coded microcarriers are initially contained in the microcarrier wells in dry form and are suspended by using a liquid handling module to transfer a sample (e.g., an antigen sample) to the microcarrier well and / or to transfer an incubation buffer from the incubation buffer well to the microcarrier well. In this case, the microcarrier well may be the reaction well.

[0065] FIG. 9 shows an exemplary immunoassay process 900 for detecting antibodies in a sample. A sample containing antibodies (in this case human IgG) is added to a reaction well containing microcarriers bound to a capture agent that contains an antigen to which the antibodies in the sample can bind (FIG. 9, "encoded microcarriers"). The magnetic unit pocket moves into the reaction well, moves up and down, and / or oscillates to promote mixing of the antibodies with the microcarriers (FIG. 7, "incubation"). The magnetic unit moves into the reaction well and into the magnetic unit pocket to attract the microcarriers to the outer surface of the magnetic unit pocket. The magnetic unit and magnetic unit pocket are transferred to a reaction wash well for washing (FIG. 7, first "wash"). The magnetic unit moves out of the reaction well to release the microcarriers into a wash buffer. The magnetic unit pocket moves up and down, and / or oscillates to promote mixing in the well. The magnetic unit moves back into the magnetic unit pocket to attract the microcarriers to the outer surface of the magnetic unit pocket. The magnetic unit and magnetic unit pocket move to the antibody well. The magnetic unit moves out of the antibody well to release the microcarriers into the antibody buffer, and the magnetic unit pocket moves, moves up and down, and / or vibrates to facilitate mixing of the microcarriers with the secondary antibody in the well (Figure 9, "Biotinylated Detection Ab Incubation"). The secondary antibody binds to the sample antigen bound to the microcarrier through its capture agent. The magnetic unit moves into the antibody well and into the magnetic unit pocket to attract the microcarriers to the outer surface of the magnetic unit pocket. The magnetic unit and magnetic unit pocket are transferred to the antibody washing well for washing (Figure 9, second "wash"). The magnetic unit moves out of the antibody washing well to release the microcarriers into the washing buffer. The magnetic unit pocket moves, moves up and down, and / or vibrates to facilitate mixing in the washing well. The magnetic unit moves back into the magnetic unit pocket to attract the microcarriers to the outer surface of the magnetic unit pocket. The magnetic unit and magnetic unit pocket move to the labeling well to transfer the microcarriers to the labeling well.The magnetic unit moves out of the labeling well to release the microcarriers into the labeling buffer, and the magnetic unit pocket moves, bobs and / or vibrates to promote mixing of the microcarriers with the labeling molecule (streptavidin-phycoerythrin; SA-PE) in the well (Figure 9, "SA-PE labeling"). The magnetic unit moves into the labeling well and into the magnetic unit pocket to attract the microcarriers to the outer surface of the magnetic unit pocket. The magnetic unit and magnetic unit pocket, along with the attracted microcarriers, are transferred to the labeling washing well for washing (Figure 9, third "wash"). After washing, the microcarriers are transferred to the detection well for optical reading of the coded microcarriers and detection of the fluorescent signal from the labeling molecule bound to the microcarriers. In some variations, the washing step is repeated in multiple washing wells.

[0066] Encoded Microcarriers The magnetic modules provided herein comprise one or more coded microcarriers, which are described in detail in International Patent Application Nos. PCT / IB2016 / 000937, PCT / US2016 / 063202 and PCT / US2022 / 031049, the entire contents of which are incorporated herein by reference.

[0067] In some aspects, a magnetic module for reacting or labeling the coded microcarriers is provided herein. In some embodiments, the coded microcarriers comprise a substantially transparent magnetic polymer layer having a first side and a second side, the first and second sides being parallel to each other, the substantially transparent magnetic polymer comprising a mixture of a substantially transparent polymer and a plurality of magnetic nanoparticles, the magnetic nanoparticles comprising iron (II, III) oxide or iron (III) oxide. In some embodiments, the coded microcarriers further comprise a substantially opaque layer, the substantially opaque layer being fixed to the first side of the substantially transparent magnetic polymer layer, and the contour of the substantially opaque layer comprises a two-dimensional shape representing the analog code. In some embodiments, the coded microcarriers further comprise one or more capture agents for capturing one or more analytes, the one or more capture agents being bound to at least one of the first side and the second side of the substantially transparent magnetic polymer layer. In some embodiments, the magnetic component of the coded microcarriers comprises a substantially transparent magnetic polymer layer. The configuration, parameters and optical characteristics of the coded microcarriers of the present disclosure are defined in the non-limiting description below.

[0068] In some embodiments, the substantially transparent magnetic polymer layer of the present disclosure comprises a mixture of a substantially transparent polymer and a plurality of magnetic nanoparticles. In some embodiments, the substantially transparent magnetic polymer layer comprises a suspension of magnetic nanoparticles in a substantially transparent polymer, such as an epoxy-based polymer. For example, the magnetic nanoparticles can be made into a stable dispersion (e.g., by using a solvent, including but not limited to, cyclopentanone or g-butyrolactone, and / or a dispersant, including but not limited to, a phosphate polymer) and mixed with the dissolved or monomeric form of the polymer. Incorporating magnetic properties into the substantially transparent polymer layer offers advantages over using a dedicated magnetic layer (e.g., a magnetic ring, etc.) by allowing a reduction in the overall size of the microcarriers, thereby increasing the number of microcarriers that can be produced from a single wafer, thereby reducing unit costs. Furthermore, the substantially transparent magnetic polymer layer does not need to be sandwiched between polymer layers to isolate it from the environment, as other dedicated magnetic layers (e.g., nickel layers) may require. This simplified two-layer model reduces manufacturing costs and increases manufacturing consistency without sacrificing functionality. Exemplary descriptions and techniques regarding magnetic nanoparticle suspensions and their mixing with epoxy-based polymers can be found, for example, in Suter, Marcel. Photopatternable superparamagnetic nanocomposite for the fabrication of microstructures. Diss. ETH Zurich, 2011; and Suter, M. (2011) Sensors and Actuators B: Chemical 156:433-43.

[0069] In some embodiments, the magnetic nanoparticles are superparamagnetic nanoparticles. Without wishing to be bound by theory, it is believed that such materials are advantageous for microcarrier fabrication because the resulting microcarriers exhibit no residual magnetism and / or only low magnetic attraction to each other (as opposed to magnetic materials such as nickel, which have some residual magnetism even after removal of the magnetic field), thus preventing them from adversely interacting in solution. Such materials are also believed to be uniquely suited for analog coding (e.g., as described herein) because the materials (e.g., SU-8 / iron (II, III) oxide or iron (III) oxide nanoparticle mixtures or suspensions, which may replace the use of nickel or rare earth metals) allow for very thin magnetic layers with improved image recognition (e.g., when imaged as described herein). These microcarriers are also believed to be simpler to fabricate. In certain embodiments, the magnetic nanoparticles are iron (III) oxide (Fe 2 O 3 : also known as ferric oxide or maghemite) and / or iron(II,III) oxide (Fe 3 O 4 : also known as magnetite. In one embodiment, the epoxy-based polymer is SU-8.

[0070] In some embodiments, the magnetic nanoparticles of the present disclosure are less than about 30 nm in diameter. In some embodiments, the magnetic nanoparticles of the present disclosure are greater than or equal to about 3 nm in diameter. In some embodiments, the magnetic nanoparticles of the present disclosure can be any size (e.g., diameter) less than about any of the following sizes: 30, 25, 20, 15, 10, or 5 (nm). In some embodiments, the magnetic nanoparticles of the present disclosure can be any size (e.g., diameter) greater than or equal to about any of the following sizes: 3, 5, 10, 15, 20, or 25 (nm). That is, the magnetic nanoparticles can be of any size (e.g., diameter) having an upper limit of 30, 25, 20, 15, 10, or 5 nm and an independently selected lower limit of 3, 5, 10, 15, 20, or 25 nm, where the lower limit is less than the upper limit.

[0071] In some embodiments, the substantially transparent magnetic polymer layer comprises a mixture of a substantially transparent polymer and a plurality of magnetic nanoparticles, the plurality of magnetic nanoparticles comprising less than about 10% (by weight) and / or more than about 0.1% (by weight) of the mixture. In some embodiments, the plurality of magnetic nanoparticles comprises about 2.5% (by weight) of the mixture. In some embodiments, the substantially transparent magnetic polymer layer of the present disclosure comprises a mixture comprising a plurality of magnetic nanoparticles at a concentration less than about any of the following concentrations: 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, or 0.3 (by weight). In some embodiments, the substantially transparent magnetic polymer layer of the present disclosure comprises a mixture comprising a plurality of magnetic nanoparticles at a concentration greater than about any of the following concentrations: 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, or 8 (by weight). That is, a substantially transparent magnetic polymer layer of the present disclosure may include a mixture including a plurality of magnetic nanoparticles in a concentration having an upper limit of 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4 or 0.3% and an independently selected lower limit of 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7 or 8%, where the lower limit is less than the upper limit.

[0072] In some embodiments, the substantially transparent polymer of the present disclosure comprises an epoxy-based polymer. Epoxy-based polymers suitable for making the compositions described herein include, but are not limited to, the EPON™ family of epoxy resins provided by Hexion Specialty Chemicals, Inc. (Columbus, Ohio) and any number of epoxy resins provided by The Dow Chemical Company (Midland, Michigan). Numerous examples of suitable polymers are commonly known in the art and include, but are not limited to, SU-8, EPON 1002F, EPON 165 / 154, and polymethylmethacrylate / polyacrylic acid block copolymers (PMMA-co-PAA). For further polymers, see, for example, Warad, IC Packaging: Package Construction Analysis in Ultra Small IC Packaging, LAP LAMBERT Academic Publishing (2010); The Electronic Packaging Handbook, CRC Press (Blackwell, ed.), (2000); and Pecht et al., Electronic Packaging Materials and Their Properties, CCR Press, 1st ed., (1998). These types of materials have the advantageous effect of not swelling in an aqueous environment, which ensures that uniform microcarrier size and shape are maintained within the population of microcarriers. In some embodiments, the substantially transparent polymer is a photoresist polymer. In some embodiments, the epoxy-based polymer is an epoxy-based negative near-ultraviolet photoresist. In some embodiments, the epoxy-based polymer is SU-8.

[0073] In some embodiments, microcarriers of the present disclosure comprise a substantially transparent magnetic polymer layer having a thickness of about 0.1 μm or more. In some embodiments, microcarriers of the present disclosure comprise a substantially transparent magnetic polymer layer having a thickness of about 50 μm or less. In some embodiments, microcarriers of the present disclosure comprise a substantially transparent magnetic polymer layer having a thickness of about 0.1 μm to about 50 μm. In some embodiments, microcarriers of the present disclosure comprise a substantially transparent magnetic polymer layer having a thickness of about no more than any of the following thicknesses: 50, 45, 40, 35, 30, 25, 20, 15, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, or 0.3 (μm). In some embodiments, microcarriers of the present disclosure comprise a substantially transparent magnetic polymer layer having a thickness of approximately any of the following thicknesses or more: 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, or 45 (μm). That is, microcarriers of the present disclosure may comprise a substantially transparent magnetic polymer layer of a thickness having an upper limit of 50, 45, 40, 35, 30, 25, 20, 15, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4 or 0.3 μm and an independently selected lower limit of 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40 or 45 μm, where the lower limit is less than the upper limit.

[0074] In some embodiments, the microcarriers of the present disclosure comprise a substantially opaque layer. For example, the substantially opaque layer may be comprised of a substantially opaque polymer or metal.

[0075] In some embodiments, the substantially opaque layer comprises a polymer described herein (e.g., SU-8) mixed with one or more opaque or colored dyes. In other embodiments, the substantially opaque layer comprises a black matrix resist. Any black matrix resist known in the art may be used. See, for example, U.S. Pat. No. 8,610,848 for exemplary black matrix resists and related methods. In some embodiments, the black matrix resist may be a photoresist colored with a black pigment, such as, for example, patterned into the color filter of an LCD as part of the black matrix. Black matrix resists may include, without limitation, those sold by Toppan Printing Co., Ltd. (now Toppan Corporation) (Tokyo), Tokyo Ohka Kogyo Co., Ltd. (Kawasaki), and Daxin Materials Corp. (Taichung, Taiwan).

[0076] In some embodiments, the substantially opaque layer comprises a substantially opaque polymer that exhibits an absorptivity greater than about 1.8 (OD) at wavelengths between about 230 nm and about 660 nm. For example, the polymer may exhibit an absorptivity of greater than about 1.8 (OD) at one or more wavelengths selected from the group consisting of 230, 240, 250, 260, 270, 280, 290, 300, 310, 320, 330, 340, 350, 360, 370, 380, 390, 400, 410, 420, 430, 440, 450, 460, 470, 480, 490, 500, 510, 520, 530, 540, 550, 560, 570, 580, 590, 600, 610, 620, 630, 640, 650, and 660 nm.

[0077] In some embodiments, the substantially opaque layer comprises a metal. In some embodiments, the metal has no remanence. In some embodiments, the substantially opaque layer comprises nickel, titanium, copper, and / or chromium.

[0078] In some embodiments, microcarriers of the present disclosure comprise a substantially opaque layer having a thickness of about 0.05 μm or more. In some embodiments, microcarriers of the present disclosure comprise a substantially opaque layer having a thickness of about 2 μm or less. In some embodiments, microcarriers of the present disclosure comprise a substantially opaque layer having a thickness of about 0.05 μm to about 2 μm. In some embodiments, microcarriers of the present disclosure comprise a substantially opaque layer having a thickness of about no more than any of the following thicknesses: 2, 1.9, 1.8, 1.7, 1.6, 1.5, 1.4, 1.3, 1.2, 1.1, 1.0, 0.9, 0.8, 0.7, 0.6, 0.5, 0.45, 0.4, 0.35, 0.3, 0.25, 0.2, 0.15, 0.125, 0.1, or 0.075 μm. In some embodiments, microcarriers of the present disclosure comprise a substantially opaque layer having a thickness of approximately any of the following thicknesses: 0.05, 0.075, 0.1, 0.125, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, or 1.8 (μm). That is, the microcarriers of the present disclosure have a particle size of 2, 1.9, 1.8, 1.7, 1.6, 1.5, 1.4, 1.3, 1.2, 1.1, 1.0, 0.9, 0.8, 0.7, 0.6, 0.5, 0.45, 0.4, 0.35, 0.3, 0.25, 0.2, 0.15, 0.125, 0.1, or 0.075 μm, where the lower limit is less than the upper limit. The substantially opaque layer may have a thickness having an upper limit and an independently selected lower limit of 0.05, 0.075, 0.1, 0.125, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7 or 1.8 μm.

[0079] In some embodiments, the analog code comprises one or more overlapping or partially overlapping arc elements that form a continuous or discontinuous ring (e.g., surrounding the center of the microcarrier). The two-dimensional shape is decoded by imaging the microcarrier (e.g., by optical microscopy) such that an image of the code is formed by the pattern produced by light passing through the substantially transparent magnetic polymer layer and light that is blocked from passing through the substantially opaque layer. Non-limiting examples of two-dimensional shapes comprised of overlapping arc elements that form discontinuous rings are shown in International Patent Application No. PCT / US2022 / 031049.

[0080] In some embodiments, microcarriers of the present disclosure may be coded with a substantially opaque layer that constitutes a two-dimensional shape. For example, as described above, the two-dimensional shape may constitute the shape of a substantially opaque layer in contact with a substantially transparent layer of the microcarrier, or may constitute the shape of the microcarrier itself (e.g., the perimeter). That is, the code is the shape of the substantially opaque layer itself (e.g., rather than a code generated by a fluorescent or other visible portion of the surface of the layer of the microcarrier). Any two-dimensional shape that can encompass multiple resolvable and characteristic variations may be used. In some embodiments, the two-dimensional shape comprises linear, circular, elliptical, rectangular, quadrilateral, or higher-angle polygonal aspects, elements, and / or shapes.

[0081] In some embodiments, the microcarrier further comprises an orientation indicator for orienting the analog code of the substantially opaque layer. Any feature of the microcarrier that is visible and / or detectable by imaging (e.g., a microscope or other forms of imaging as described herein) and / or by image recognition software may act as an orientation indicator. The orientation indicator may act as a reference point for, for example, an image recognition algorithm to orient the image of the analog code in a uniform orientation (i.e., the shape of the substantially opaque layer). This advantageously simplifies image recognition, as the algorithm only needs to compare an image of a particular analog code against a library of analog codes in the same orientation, and not against a library comprising all analog codes in all possible orientations. In some embodiments, the orientation indicator comprises an asymmetry of the substantially opaque layer, such as a discontinuity in its profile or shape. For example, the orientation indicator may comprise a visible feature, such as an asymmetry in the two-dimensional shape representing the analog code of the microcarrier.

[0082] In some embodiments, the microcarriers of the present disclosure are generally circular disks. As used herein, a generally circular shape can refer to any shape in which the distance between all of the points on the perimeter of the shape and the geometric center of the shape is approximately the same. In some embodiments, a shape is considered to be generally circular if the variation between any of the potential radii connecting the geometric center and a given point on the perimeter varies by no more than 10% in length. As used herein, a generally circular disk can refer to any generally circular shape in which the thickness of the shape is significantly less than its diameter. For example, in some embodiments, the thickness of a generally circular disk can be less than about 50%, less than about 40%, less than about 30%, less than about 20%, less than about 15%, less than about 10%, or less than about 5% of its diameter. In some embodiments, the thickness of a generally circular disk can be about 20% of its diameter.

[0083] In some embodiments, the microcarriers are less than about 200 μm in diameter. For example, in some embodiments, the microcarriers are less than about 200 μm, less than about 180 μm, less than about 160 μm, less than about 140 μm, less than about 120 μm, less than about 100 μm, less than about 80 μm, less than about 60 μm, less than about 40 μm, or less than about 20 μm in diameter. In some embodiments, the microcarriers are more than about 5 μm, more than about 10 μm, more than about 20 μm, more than about 30 μm, more than about 40 μm, more than about 50 μm, more than about 60 μm, more than about 70 μm, more than about 80 μm, more than about 90 μm, more than about 100 μm, more than about 120 μm, more than about 140 μm, or more than about 150 μm in diameter. In some embodiments, microcarriers of the present disclosure can be any size (e.g., diameter) less than about any of the following sizes: 200, 180, 160, 140, 120, 100, 80, 60, 40, or 20 (μm). In some embodiments, magnetic nanoparticles of the present disclosure can be any size (e.g., diameter) greater than or equal to about any of the following sizes: 5, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 120, 140, or 150 (μm). That is, the magnetic nanoparticles can be of any size (e.g., diameter) having an upper limit of 200, 180, 160, 140, 120, 100, 80, 60, 40, or 20 μm and an independently selected lower limit of 5, 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 120, 140, or 150 μm, where the lower limit is less than the upper limit.

[0084] In some embodiments, the microcarriers have a diameter of about 180 μm, about 160 μm, about 140 μm, about 120 μm, about 100 μm, about 90 μm, about 80 μm, about 70 μm, about 60 μm, about 50 μm, about 40 μm, about 30 μm, about 20 μm, or about 10 μm. In some embodiments, the microcarriers are about 40 μm in diameter.

[0085] In some embodiments, the microcarriers are less than about 50 μm thick. For example, in some embodiments, the microcarriers are less than about 70 μm, less than about 60 μm, less than about 50 μm, less than about 40 μm, less than about 30 μm, less than about 25 μm, less than about 20 μm, less than about 15 μm, less than about 10 μm, or less than about 5 μm thick. In some embodiments, the microcarriers are less than about any of the following thicknesses: 50, 45, 40, 35, 30, 25, 20, 15, 10, 9, 8, 7, 6, 5, 4, 3, or 2 μm. In some embodiments, the microcarriers are more than about the following thicknesses: 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, or 45 μm. That is, the thickness of the microcarrier can be any of a range of thicknesses having upper limits of 50, 45, 40, 35, 30, 25, 20, 15, 10, 9, 8, 7, 6, 5, 4, 3 or 2 and independently selected lower limits of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40 or 45 (μm), where the lower limit is less than the upper limit.

[0086] In some embodiments, the microcarrier has a thickness of about 50 μm, about 45 μm, about 40 μm, about 35 μm, about 30 μm, about 25 μm, about 20 μm, about 19 μm, about 18 μm, about 17 μm, about 16 μm, about 15 μm, about 14 μm, about 13 μm, about 12 μm, about 11 μm, about 10 μm, about 9 μm, about 8 μm, about 7 μm, about 6 μm, about 5 μm, about 4 μm, about 3 μm, about 2 μm, or about 1 μm. In some embodiments, the microcarrier has a thickness of about 50 μm to about 2 μm, about 20 μm to about 2 μm, or about 10 μm to about 2 μm. In certain embodiments, the microcarrier is about 5 μm thick.

[0087] III. Assay Systems Certain aspects of the present disclosure relate to an assay system comprising any of the magnetic modules provided herein. In some embodiments, the assay system further comprises a detection module. In some embodiments, the detection module is configured to detect a signal from the magnetic microcarriers. Any type of detectable signal may be used, such as fluorescence intensity, anisotropy, luminescence, color, visible pattern, concentration, reaction product, nucleotide sequence, and polypeptide sequence. In some embodiments, the signal is a fluorescent signal.

[0088] In some embodiments, the assay system comprises one or more detection wells. In some embodiments, the one or more detection wells are positioned in the same one-dimensional or two-dimensional array as the various wells in the magnetic module. In some embodiments, the one or more detection wells are positioned in the same one-dimensional or two-dimensional array as the final label wash well such that microcarriers in the final label wash well can be transferred to one or more detection wells simultaneously.

[0089] The one or more detection wells may be of any suitable shape and material. In some embodiments, the one or more detection wells have a flat bottom. In some embodiments, the one or more detection wells are substantially transparent. In some embodiments, the one or more detection wells are substantially opaque.

[0090] In some embodiments, the one or more detection wells and the one or more magnetic units have the same one-dimensional or two-dimensional arrangement, such that one or more magnetic units can move into one or more detection wells at the same time and out of one or more detection wells at the same time. In some embodiments, the one or more detection wells and the one or more magnetic unit pockets have the same one-dimensional or two-dimensional arrangement, such that one or more magnetic unit pockets can move into one or more detection wells at the same time and out of one or more detection wells at the same time.

[0091] In some embodiments, the assay system further comprises a PCR module. Any PCR module configured to perform a polymerase chain reaction (PCR) may be used. In some embodiments, the PCR module is configured to perform a PCR using primers bound to one or more secondary binding moieties capable of binding to a target molecule. In some embodiments, the secondary binding moiety comprises biotin.

[0092] In some embodiments, the assay system further comprises a liquid handling module. The liquid handling module is configured to transfer liquid to and from any location within or outside the assay system. For example, the liquid handling module can transfer liquid (e.g., sample or buffer) to, from and between wells of a magnetic module. In some embodiments, the liquid handling module is configured to transfer sample (e.g., amplicon) from a PCR module to a magnetic module (e.g., a reaction well or a microcarrier well).

[0093] In some embodiments, the assay system further comprises a hybridization buffer well. In some embodiments, a hybridization buffer well is in each well set. In some embodiments, the hybridization buffer well comprises a hybridization buffer. In some embodiments, the liquid handling module is configured to transfer at least a portion of the hybridization buffer from the hybridization buffer well to the reaction well or the microcarrier well.

[0094] In some embodiments, the assay system further comprises incubation buffer wells. In some embodiments, an incubation buffer well is in each well set. In some embodiments, the incubation buffer wells comprise an incubation buffer. In some embodiments, the liquid handling module is configured to transfer at least a portion of the incubation buffer from the incubation buffer wells to the reaction wells or the microcarrier wells.

[0095] IV. METHODS OF CONDUCTING ASSAYS USING THE MAGNETIC MODULE AND ASSAY SYSTEM Certain aspects of the present disclosure relate to methods of reacting or labeling magnetic microcarriers and using the magnetic modules described herein to detect target molecules or perform multiplex assays such as immunoassays. Advantageously, the magnetic modules allow for assays with increased throughput and reduced recognition errors using a large number of potentially unique microcarriers.

[0096] In some embodiments, the method comprises optically reading the magnetic microcarriers. In some embodiments, optically reading the magnetic microcarriers uses analog shape recognition to decode the analog code to identify the microcarriers. Conceptually, this decoding may involve imaging the analog code of each microcarrier (e.g., in a solution or sample), comparing each image to a library of analog codes, and matching each image with an image from the library, thereby positively identifying the code. Optionally, as described herein, when using microcarriers with an orientation indicator (e.g., asymmetry), the decoding may further comprise rotating each image to align it to a particular orientation (e.g., based in part on the orientation indicator). For example, when the orientation indicator comprises a space, the image may be rotated until the space reaches a predetermined position or orientation (e.g., the 0° position of the image).

[0097] Various shape recognition software, tools and methods are known in the art. Examples of such APIs and tools include, but are not limited to, Microsoft® Research FaceSDK, OpenBR, Face and Scene Recognition by ReKognition, Betaface API and various ImageJ plug-ins. In some embodiments, analog shape recognition may comprise image processing steps such as, but not limited to, foreground extraction, shape detection, thresholding (e.g., automatic image thresholding or manual image thresholding).

[0098] One of skill in the art will appreciate that the methods and microcarriers described herein may be adapted for a variety of imaging devices, including, without limitation, microscopes, plate readers, etc. In some embodiments, decoding the analog code may comprise illuminating the microcarrier by passing light through a substantially transparent portion of the microcarrier (e.g., a substantially transparent polymer layer) and / or a surrounding solution. The light may then be unable to penetrate, or may penetrate with a lower intensity or other significant difference, a substantially opaque portion of the microcarrier (e.g., a substantially opaque polymer layer), producing an analog coded light pattern corresponding to the microcarrier.

[0099] As mentioned above, any type of optical microscopy may be used in the methods of the present disclosure, including, without limitation, one or more of bright field microscopy, dark field microscopy, phase contrast microscopy, differential interference contrast microscopy (DIC), Nomarski interference microscopy (NIC), Nomarski microscopy, Hoffman modulation contrast microscopy (HMC), or fluorescence microscopy. In certain embodiments, the analog code may be decoded using bright field microscopy and the analyte may be detected using fluorescence microscopy.

[0100] Methods for molecular assays In some embodiments, provided herein are methods for performing or facilitating molecular assays using the magnetic modules or assay systems provided herein. In some embodiments, the methods comprise (a) transferring one or more analytes into a reaction well, the reaction well containing magnetic microcarriers in a reaction buffer. In some embodiments, the methods comprise (b) moving one of the one or more magnetic unit pockets into the reaction well and moving or vibrating the magnetic unit pocket within the reaction well to mix the one or more analytes with the magnetic microcarriers. In some embodiments, the methods comprise (c) moving the magnetic unit into the reaction well and into the magnetic unit pocket to attract the magnetic microcarriers to an outer surface of the magnetic unit pocket. In some embodiments, the methods comprise (d) transferring the magnetic microcarriers from the reaction well to a first reaction wash well by moving the magnetic unit pocket and magnetic unit out of the reaction well and into a first reaction wash well. In some embodiments, the methods comprise (e) transferring the magnetic microcarriers from a final reaction wash well to a label well by moving the magnetic unit pocket and magnetic unit out of the final reaction wash well and into a label well. In some embodiments, the method comprises (f) moving the magnetic unit out of the labeling well while retaining the magnetic unit pocket within the labeling well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the labeling buffer. In some embodiments, the method comprises (g) moving the magnetic unit into the labeling well and into the magnetic unit pocket to attract the magnetic microcarriers to the outer surface of the magnetic unit pocket. In some embodiments, the method comprises (h) moving the magnetic unit pocket and magnetic unit out of the labeling well and into a first label washing well, thereby transferring the magnetic microcarriers from the labeling well to the first label washing well.

[0101] It should be appreciated that steps (a)-(h) described in the above paragraphs may be performed in any order, and any step may be optionally repeated or omitted, as desired. In some embodiments, steps (a)-(h) are performed in the order listed above. In some embodiments, a mixing step similar to step (b) is performed after transferring and releasing the microcarriers into each well to facilitate mixing of the microcarriers with the buffer or reagent in each well. In some embodiments, any of the washing steps may be optionally repeated.

[0102] In some embodiments, the method further comprises, after step (d) and before step (e), moving the magnetic unit out of the first reaction wash well while retaining the magnetic unit pocket within the first reaction wash well, thereby releasing the magnetic microcarriers from an outer surface of the magnetic unit pocket into the first reaction wash well. In some embodiments, the method further comprises, after step (d) and before step (e), moving or vibrating the magnetic unit pocket within the first reaction wash well to mix the magnetic microcarriers with the reaction wash buffer.

[0103] In some embodiments, the method further comprises, after step (f) and before step (g), moving or vibrating the magnetic unit pocket within the label well to mix the magnetic microcarriers with the labeling buffer. In some embodiments, the method further comprises, after step (h), moving the magnetic unit out of the first labeling wash well while retaining the magnetic unit pocket within the first labeling wash well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the first labeling wash well. In some embodiments, the method further comprises, after step (h), moving or vibrating the magnetic unit pocket within the first labeling wash well to mix the magnetic microcarriers with the labeling wash buffer.

[0104] In some embodiments, the method further comprises, after step (h), transferring the magnetic microcarriers from the final reaction washing well to the detection well by moving the magnetic unit pocket and the magnetic unit out of the final labeling washing well and into the detection well.

[0105] In some embodiments, the method further comprises the step of moving the magnetic unit out of the detection well while retaining the magnetic unit pocket within the detection well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the detection well.

[0106] In some embodiments, the method further comprises, after step (h), optically reading the magnetic microcarriers to detect signals from labelled molecules bound to the magnetic microcarriers.

[0107] Methods for Immunoassays In some embodiments, provided herein are methods for performing molecular assays using the magnetic modules or assay systems provided herein. In some embodiments, the methods comprise (a) transferring one or more analytes into a reaction well, the reaction well containing magnetic microcarriers in a reaction buffer. In some embodiments, the methods comprise (b) moving one of the one or more magnetic unit pockets into the reaction well and moving or vibrating the magnetic unit pocket within the reaction well to mix the one or more analytes with the magnetic microcarriers. In some embodiments, the methods comprise (c) moving the magnetic unit into the reaction well and into the magnetic unit pocket to attract the magnetic microcarriers to an outer surface of the magnetic unit pocket. In some embodiments, the methods comprise (d) transferring the magnetic microcarriers from the reaction well to a first reaction wash well by moving the magnetic unit pocket and magnetic unit out of the reaction well and into a first reaction wash well. In some embodiments, the methods comprise (e) transferring the magnetic microcarriers from a final reaction wash well to an antibody well by moving the magnetic unit pocket and magnetic unit out of the final reaction wash well and into an antibody well. In some embodiments, the method comprises (f) moving the magnetic unit out of the antibody well while retaining the magnetic unit pocket within the label well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the antibody buffer. In some embodiments, the method comprises (g) moving the magnetic unit into the antibody well and into the magnetic unit pocket to attract the magnetic microcarriers to the outer surface of the magnetic unit pocket. In some embodiments, the method comprises (h) transferring the magnetic microcarriers from the antibody well to the first antibody washing well by moving the magnetic unit pocket and magnetic unit out of the antibody well and into the first antibody washing well. In some embodiments, the method comprises (i) transferring the magnetic microcarriers from the final antibody washing well to the label well by moving the magnetic unit pocket and magnetic unit out of the final antibody washing well and into the label well.In some embodiments, the method comprises (j) moving the magnetic unit out of the labeling well while retaining the magnetic unit pocket within the labeling well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the labeling buffer. In some embodiments, the method comprises (k) moving the magnetic unit into the labeling well and into the magnetic unit pocket to attract the magnetic microcarriers to the outer surface of the magnetic unit pocket. In some embodiments, the method comprises (l) transferring the magnetic microcarriers from the labeling well to the first labeling washing well by moving the magnetic unit pocket and magnetic unit out of the labeling well and into a first labeling washing well.

[0108] It should be appreciated that steps (a)-(l) described in the above paragraphs may be performed in any order, and any step may be optionally repeated or omitted, as desired. In some embodiments, steps (a)-(l) are performed in the order listed. In some embodiments, a mixing step similar to step (b) is performed after transferring and releasing the microcarriers into each well to facilitate mixing of the microcarriers with the buffer or reagent in each well. In some embodiments, any of the washing steps may be optionally repeated.

[0109] In some embodiments, the method further comprises, after step (d) and before step (e), moving the magnetic unit out of the first reaction wash well while retaining the magnetic unit pocket within the first reaction wash well, thereby releasing the magnetic microcarriers from an outer surface of the magnetic unit pocket into the first reaction wash well. In some embodiments, the method further comprises, after step (d) and before step (e), moving or vibrating the magnetic unit pocket within the first reaction wash well to mix the magnetic microcarriers with the reaction wash buffer.

[0110] In some embodiments, the method further comprises, after step (f) and before step (g), moving or vibrating the magnetic unit pocket within the antibody well to mix the magnetic microcarriers with the labelling buffer.

[0111] In some embodiments, the method further comprises, after step (h) and before step (i), moving the magnetic unit out of the first antibody washing well while retaining the magnetic unit pocket within the first antibody washing well, thereby releasing the magnetic microcarriers from an outer surface of the magnetic unit pocket into the first antibody washing well. In some embodiments, the method further comprises, after step (h) and before step (i), moving or vibrating the magnetic unit pocket within the first antibody washing well to mix the magnetic microcarriers with the antibody washing buffer.

[0112] In some embodiments, the method further comprises, after step (j) and before step (k), moving or vibrating the magnetic unit pocket within the labelling well to mix the magnetic microcarriers with the labelling buffer.

[0113] In some embodiments, the method further comprises, after step (l), moving the magnetic unit out of the first label wash well while retaining the magnetic unit pocket within the first label wash well, thereby releasing the magnetic microcarriers from an outer surface of the magnetic unit pocket into the first label wash well. In some embodiments, the method further comprises, after step (l), moving or vibrating the magnetic unit pocket within the first label wash well to mix the magnetic microcarriers with the label wash buffer.

[0114] In some embodiments, the method further comprises, after step (l), transferring the magnetic microcarriers from the final reaction washing well to the detection well by moving the magnetic unit pocket and magnetic unit out of the final labeling washing well and into the detection well.

[0115] In some embodiments, the method further comprises the step of moving the magnetic unit out of the detection well while retaining the magnetic unit pocket within the detection well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the detection well.

[0116] In some embodiments, the method further comprises, after step (l), optically reading the magnetic microcarriers to detect signals from labelled molecules bound to the magnetic microcarriers. EXAMPLES

[0117] The presently disclosed subject matter will be better understood by reference to the following examples, which are provided by way of illustration of the invention and not by way of limitation.

[0118] Example 1: Exemplary molecular assays using the magnetic module Exemplary molecular assays were performed using the magnetic module described herein. Control molecular assay experiments using conventional methods were performed, where coded microcarriers were processed in a single well using dispense and aspiration channels to add or remove buffers in the well and a shaking platform to mix the microcarriers with different buffers. Median fluorescence intensity (MFI) was measured for molecular assays and control experiments using the magnetic module described herein. Background signal was used to monitor the removal of excess SA-PE through the washing process. The results are summarized in Table 1. The small difference in the mean signal MFI of less than 1.3%, calculated as (107724-106377) / 106377, and the small difference in background signal of less than 0.13%, calculated as (1596-1594) / 1596, indicate that the performance of the magnetic module is comparable to that of the conventional single-well method, while achieving greater simplicity in processing and device design. Table 1. Molecular assay results using the magnetic module and single-well methods. [Table 1]

[0119] Example 2: Exemplary Immunoassay Using the Magnetic Module An exemplary immunoassay was performed using the magnetic module described herein. Median fluorescence intensity (MFI) was measured for the immunoassay at various antigen (pregnancy associated plasma protein A, PAPP-A) concentrations. The results are summarized in Table 2 below and FIG. 10. The results show that the assay system using the magnetic module can generate higher signals with increasing antigen concentrations. The similar MFI between experiment 1 and experiment 2 at the same antigen concentrations indicates that the assay system using the magnetic module is valid and stable. Table 2. Immunoassay results using the magnetic module [Table 2]

Claims

1. A magnetic module for reacting or labeling magnetic microcarriers, comprising: one or more well sets, each well set comprising: A reaction well; one or more reaction wash wells comprising a first reaction wash well and a final reaction wash well, each reaction wash well containing a reaction wash buffer, and when there is only one reaction wash well in the well set, the first reaction wash well and the final reaction wash well are the same well; a labeling well containing a labeling molecule in a labeling buffer; one or more label wash wells comprising a first label wash well and a final label wash well, each label wash well containing a label wash buffer, and when there is only one label wash well in the well set, the first label wash well and the final label wash well are the same well; one or more well sets comprising: One or more magnetic units, each magnetic unit comprising: configured to move into, out of, and between each of the wells in the well set; configured to attract the magnetic microcarriers and move the magnetic microcarriers into, out of, and between each of the wells in the well set; having an upper end and a lower end; one or more magnetic units; One or more magnetic unit pockets, each magnetic unit pocket comprising: Positioned between each magnetic unit and the well set, configured to move into, out of, and between each of the wells in the well set; configured to at least partially cover the bottom end of each magnetic unit as the magnetic unit moves or slides into the magnetic unit pocket; having an inner surface and an outer surface; one or more magnetic unit pockets; Equipped with A magnetic module, wherein the magnetic microcarriers are coupled to one or more capture agents, the one or more capture agents being capable of binding to one or more analytes.

2. Each well set is an antibody well containing a secondary antibody in an antibody buffer, said secondary antibody being capable of binding to said one or more analytes and being bound to one or more secondary binding moieties; one or more antibody wash wells comprising a first antibody wash well and a final antibody wash well, each antibody wash well containing an antibody wash buffer, and when there is only one antibody wash well in the well set, the first antibody wash well and the final antibody wash well are the same well; The magnetic module of claim 1 further comprising:

3. 3. The magnetic module of claim 2, wherein the reaction well, the one or more reaction wash wells, the antibody well, the one or more antibody wash wells, the label well and the one or more label wash wells are positioned in that order along a straight line or a curved line.

4. The magnetic module of claim 1 , wherein the reaction well, the one or more reaction wash wells, the label well and the one or more label wash wells are positioned in that order along a straight line or a curved line.

5. 2. The magnetic module of claim 1, wherein each of the one or more capture agents is independently selected from the group consisting of small molecules, polynucleotides, polypeptides, proteins, lipids and polysaccharides, and each of the one or more analytes is independently selected from the group consisting of small molecules, polynucleotides, polypeptides, proteins, lipids and polysaccharides.

6. each of the one or more capture agents and the one or more analytes is independently selected from the group consisting of polynucleotides; each of the one or more capture agents is independently selected from the group consisting of antibodies and antibody fragments, and each of the one or more analytes is independently selected from the group consisting of antigens; or The magnetic module of claim 1 , wherein each of said one or more capture agents is independently selected from the group consisting of antigens and each of said one or more analytes is independently selected from the group consisting of antibodies.

7. The magnetic module of claim 1 , wherein the label molecule comprises a label binding moiety bound to a report moiety.

8. 8. The magnetic module of claim 7, wherein the label binding moiety is streptavidin (SA) or an anti-biotin antibody and the report moiety is phycoerythrin (PE) or fluorescein isothiocyanate (FITC).

9. The magnetic module according to claim 1 , wherein each magnetic unit has a rod-like shape, and each magnetic unit pocket has a bottomed cylindrical shape.

10. The magnetic module of claim 1 , wherein the one or more magnetic unit pockets are configured to move or oscillate vertically relative to the one or more well sets.

11. The magnetic module according to claim 1 , wherein the number of said one or more magnetic units, the number of said one or more magnetic unit pockets and the number of said reaction wells are the same.

12. 2. The magnetic module of claim 1, wherein the one or more sets of wells form a multiwell plate, each set of wells forming a respective row of the multiwell plate, and each type of well forming a respective column of the multiwell plate.

13. The magnetic module of claim 12, wherein the one or more magnetic units are positioned above the multiwell plate along a straight line and parallel to the row direction of the multiwell plate, and the one or more magnetic unit pockets are positioned between the multiwell plate and the one or more magnetic units along a straight line and parallel to the row direction of the multiwell plate.

14. The magnetic module of claim 1 , wherein the reaction wells are positioned along a surface or form a two-dimensional array, or the label wells are positioned along a surface or form a two-dimensional array, or both.

15. 10. The magnetic module of claim 1, wherein the various wells form a multiwell plate such that the one or more sets of wells comprise a plurality of the multiwell plate.

16. 2. The magnetic module of claim 1, wherein the various wells are positioned along a line or form a one-dimensional array, and the one or more magnetic units, the one or more magnetic unit pockets and the various wells are in the same one-dimensional or two-dimensional array, such that the one or more magnetic units can move into and out of the various wells at the same time, and the one or more magnetic unit pockets can move into and out of the various wells at the same time.

17. The magnetic module of claim 1 , wherein the magnetic microcarriers comprise encoded microcarriers, spherical magnetic beads, or a combination thereof.

18. The magnetic microcarriers include encoded microcarriers, each of the encoded microcarriers comprising: (a) a substantially transparent magnetic polymer layer having a first surface and a second surface, the first and second surfaces being parallel to one another, the substantially transparent magnetic polymer comprising a mixture of a substantially transparent polymer and a plurality of magnetic nanoparticles, the magnetic nanoparticles comprising iron (II, III) oxide or iron (III) oxide; (b) a substantially opaque layer secured to the first surface of the substantially transparent magnetic polymer layer, the substantially opaque layer having a contour that defines a two-dimensional shape that represents an analog code; and (c) one or more capture agents for capturing one or more analytes, the one or more capture agents being bound to at least one of the first surface and the second surface of the substantially transparent magnetic polymer layer; and The magnetic module of claim 1 , comprising:

19. The magnetic module of claim 18 , wherein the magnetic nanoparticles are superparamagnetic.

20. 20. The magnetic module of claim 18, wherein the microcarriers are between about 5 μm and about 200 μm in diameter, and the microcarriers are about 40 μm in diameter.

21. 20. The magnetic module of claim 18, wherein each of the one or more analytes and the one or more capture agents is independently selected from the group consisting of DNA molecules, DNA-like molecules, RNA molecules, RNA-like molecules, polynucleotides, proteins, enzymes, lipids, phospholipids, carbohydrate moieties, polysaccharides, antigens, viruses, cells, antibodies, small molecules, bacterial cells, organelles, and antibody fragments.

22. An assay system comprising the magnetic module of claim 1 .

23. 23. The assay system of claim 22, further comprising a detection module configured to detect a signal from the magnetic microcarriers.

24. 24. The assay system of claim 23, wherein the detection module comprises one or more detection wells, the one or more detection wells comprising the final label wash well.

25. The assay system of claim 23, wherein the detection module comprises one or more detection wells, and the one or more detection wells, the one or more magnetic units and the one or more magnetic unit pockets have the same one-dimensional or two-dimensional array such that the one or more magnetic units can move into the one or more detection wells and out of the one or more detection wells simultaneously, and the one or more magnetic unit pockets can move into the one or more detection wells and out of the one or more detection wells simultaneously.

26. 23. The assay system of claim 22, further comprising a polymerase chain reaction (PCR) module configured to perform a PCR.

27. 23. The assay system of claim 22, further comprising a liquid handling module configured to transfer sample from the PCR module to the reaction well of the magnetic module.

28. 28. The assay system of claim 27, wherein each well set further comprises a hybridization buffer well containing a hybridization buffer, and wherein the liquid handling module is configured to transfer at least a portion of the hybridization buffer from the hybridization buffer wells to the reaction wells.

29. 28. The assay system of claim 27, wherein each well set further comprises an incubation buffer well containing an incubation buffer, and wherein the liquid handling module is configured to transfer at least a portion of the incubation buffer from the incubation buffer well to the reaction well.

30. 2. A method of using the magnetic module of claim 1, comprising: (a) transferring one or more analytes into the reaction well, the reaction well containing the magnetic microcarriers in a reaction buffer; (b) moving one of the one or more magnetic unit pockets into the reaction well and moving or vibrating the magnetic unit pocket within the reaction well to mix the one or more analytes with the magnetic microcarriers; (c) moving the magnetic unit into the reaction well and into the magnetic unit pocket to attract the magnetic microcarriers to an outer surface of the magnetic unit pocket; (d) transferring the magnetic microcarriers from the reaction well to the first reaction-wash well by moving the magnetic unit pocket and magnetic unit out of the reaction well and into the first reaction-wash well; (e) transferring the magnetic microcarriers from the final reaction washing well to the labeling well by moving the magnetic unit pocket and magnetic unit out of the final reaction washing well and into the labeling well; (f) moving the magnetic unit out of the labeling well while retaining the magnetic unit pocket within the labeling well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the labeling buffer; (g) moving the magnetic unit into the labeling well and into the magnetic unit pocket to attract the magnetic microcarriers to an outer surface of the magnetic unit pocket; (h) transferring the magnetic microcarriers from the label well to the first label washing well by moving the magnetic unit pocket and magnetic unit out of the label well and into the first label washing well; A method for providing the above.

31. 31. The method of claim 30, wherein steps (a) through (h) are performed in the order recited.

32. After step (d) and before step (e), moving the magnetic unit out of the first reaction-wash well while holding the magnetic unit pocket within the first reaction-wash well, thereby releasing the magnetic microcarriers from an outer surface of the magnetic unit pocket into the first reaction-wash well; moving or vibrating the magnetic unit pocket within the first reaction-wash well to mix the magnetic microcarriers with the reaction-wash buffer; 31. The method of claim 30 further comprising:

33. 31. The method of claim 30, further comprising the step of moving or vibrating the magnetic unit pocket within the labeling well after step (f) and before step (g) to mix the magnetic microcarriers with the labeling buffer.

34. After step (h), moving the magnetic unit out of the first label wash well while holding the magnetic unit pocket within the first label wash well, thereby releasing the magnetic microcarriers from an outer surface of the magnetic unit pocket into the first label wash well; moving or vibrating the magnetic unit pocket within the first label wash well to mix the magnetic microcarriers with the label wash buffer; 31. The method of claim 30 further comprising:

35. The method of claim 30, further comprising, after step (h), a step of transferring the magnetic microcarriers from the final reaction washing well to the detection well by moving the magnetic unit pocket and magnetic unit out of the final labeling washing well and into the detection well.

36. The method of claim 35, further comprising the step of moving the magnetic unit out of the detection well while retaining the magnetic unit pocket within the detection well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the detection well.

37. 31. The method of claim 30, further comprising the step of optically reading the magnetic microcarriers after step (h) to detect a signal from a label molecule bound to the magnetic microcarriers.

38. A method of using the magnetic module according to claim 2, comprising the steps of: (a) transferring one or more analytes into the reaction well, the reaction well containing the magnetic microcarriers in a reaction buffer; (b) moving one of the one or more magnetic unit pockets into the reaction well and moving or vibrating the magnetic unit pocket within the reaction well to mix the one or more analytes with the magnetic microcarriers; (c) moving the magnetic unit into the reaction well and into the magnetic unit pocket to attract the magnetic microcarriers to an outer surface of the magnetic unit pocket; (d) transferring the magnetic microcarriers from the reaction well to the first reaction-wash well by moving the magnetic unit pocket and magnetic unit out of the reaction well and into the first reaction-wash well; (e) transferring the magnetic microcarriers from the final reaction washing well to the antibody well by moving the magnetic unit pocket and magnetic unit out of the final reaction washing well and into the antibody well; (f) moving the magnetic unit out of the antibody well while maintaining the magnetic unit pocket within the label well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the antibody buffer; (g) moving the magnetic unit into the antibody well and into the magnetic unit pocket to attract the magnetic microcarriers to the outer surface of the magnetic unit pocket; (h) transferring the magnetic microcarriers from the antibody well to the first antibody washing well by moving the magnetic unit pocket and magnetic unit out of the antibody well and into the first antibody washing well; (i) transferring the magnetic microcarriers from the final antibody washing well to the labeling well by moving the magnetic unit pocket and magnetic unit out of the final antibody washing well and into the labeling well; (j) moving the magnetic unit out of the labeling well while retaining the magnetic unit pocket within the labeling well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the labeling buffer; (k) moving the magnetic unit into the labeling well and the magnetic unit pocket to attract the magnetic microcarriers to an outer surface of the magnetic unit pocket; (l) transferring the magnetic microcarriers from the label well to the first label washing well by moving the magnetic unit pocket and magnetic unit out of the label well and into the first label washing well; A method for providing the above.

39. 40. The method of claim 38, wherein steps (a) through (l) are performed in the order recited.

40. After step (d) and before step (e), moving the magnetic unit out of the first reaction-wash well while holding the magnetic unit pocket within the first reaction-wash well, thereby releasing the magnetic microcarriers from an outer surface of the magnetic unit pocket into the first reaction-wash well; moving or vibrating the magnetic unit pocket within the first reaction-wash well to mix the magnetic microcarriers with the reaction-wash buffer; 40. The method of claim 38, further comprising:

41. 39. The method of claim 38, further comprising the step of moving or vibrating the magnetic unit pocket within the antibody well after step (f) and before step (g) to mix the magnetic microcarriers with the labeling buffer.

42. After step (h) and before step (i), moving the magnetic unit out of the first antibody washing well while retaining the magnetic unit pocket within the first antibody washing well, thereby releasing the magnetic microcarriers from an outer surface of the magnetic unit pocket into the first antibody washing well; moving or vibrating the magnetic unit pocket within the first antibody wash well to mix the magnetic microcarriers with the antibody wash buffer; 40. The method of claim 38, further comprising:

43. 39. The method of claim 38, further comprising the step of moving or vibrating the magnetic unit pocket within the labeling well after step (j) and before step (k) to mix the magnetic microcarriers with the labeling buffer.

44. After step (l), moving the magnetic unit out of the first label wash well while holding the magnetic unit pocket within the first label wash well, thereby releasing the magnetic microcarriers from an outer surface of the magnetic unit pocket into the first label wash well; moving or vibrating the magnetic unit pocket within the first label wash well to mix the magnetic microcarriers with the label wash buffer; 40. The method of claim 38, further comprising:

45. The method of claim 38, further comprising, after step (l), a step of transferring the magnetic microcarriers from the final reaction washing well to the detection well by moving the magnetic unit pocket and magnetic unit out of the final labeling washing well and into the detection well.

46. The method of claim 45, further comprising the step of moving the magnetic unit out of the detection well while retaining the magnetic unit pocket within the detection well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the detection well.

47. 40. The method of claim 38, further comprising the step of optically reading the magnetic microcarriers after step (l) to detect a signal from a label molecule bound to the magnetic microcarriers.

48. 13. A method of using an assay system comprising the magnetic module of claim 1, comprising: (a) transferring one or more analytes into the reaction well, the reaction well containing the magnetic microcarriers in a reaction buffer; (b) moving one of the one or more magnetic unit pockets into the reaction well and moving or vibrating the magnetic unit pocket within the reaction well to mix the one or more analytes with the magnetic microcarriers; (c) moving the magnetic unit into the reaction well and into the magnetic unit pocket to attract the magnetic microcarriers to an outer surface of the magnetic unit pocket; (d) transferring the magnetic microcarriers from the reaction well to the first reaction-wash well by moving the magnetic unit pocket and magnetic unit out of the reaction well and into the first reaction-wash well; (e) transferring the magnetic microcarriers from the final reaction washing well to the labeling well by moving the magnetic unit pocket and magnetic unit out of the final reaction washing well and into the labeling well; (f) moving the magnetic unit out of the labeling well while retaining the magnetic unit pocket within the labeling well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the labeling buffer; (g) moving the magnetic unit into the labeling well and into the magnetic unit pocket to attract the magnetic microcarriers to an outer surface of the magnetic unit pocket; (h) transferring the magnetic microcarriers from the label well to the first label washing well by moving the magnetic unit pocket and magnetic unit out of the label well and into the first label washing well; A method for providing the above.

49. A method of using an assay system comprising the magnetic module of claim 2, comprising: (a) transferring one or more analytes into the reaction well, the reaction well containing the magnetic microcarriers in a reaction buffer; (b) moving one of the one or more magnetic unit pockets into the reaction well and moving or vibrating the magnetic unit pocket within the reaction well to mix the one or more analytes with the magnetic microcarriers; (c) moving the magnetic unit into the reaction well and into the magnetic unit pocket to attract the magnetic microcarriers to an outer surface of the magnetic unit pocket; (d) transferring the magnetic microcarriers from the reaction well to the first reaction-wash well by moving the magnetic unit pocket and magnetic unit out of the reaction well and into the first reaction-wash well; (e) transferring the magnetic microcarriers from the final reaction washing well to the antibody well by moving the magnetic unit pocket and magnetic unit out of the final reaction washing well and into the antibody well; (f) moving the magnetic unit out of the antibody well while maintaining the magnetic unit pocket within the label well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the antibody buffer; (g) moving the magnetic unit into the antibody well and into the magnetic unit pocket to attract the magnetic microcarriers to the outer surface of the magnetic unit pocket; (h) transferring the magnetic microcarriers from the antibody well to the first antibody washing well by moving the magnetic unit pocket and magnetic unit out of the antibody well and into the first antibody washing well; (i) transferring the magnetic microcarriers from the final antibody washing well to the labeling well by moving the magnetic unit pocket and magnetic unit out of the final antibody washing well and into the labeling well; (j) moving the magnetic unit out of the labeling well while retaining the magnetic unit pocket within the labeling well, thereby releasing the magnetic microcarriers from the outer surface of the magnetic unit pocket into the labeling buffer; (k) moving the magnetic unit into the labeling well and into the magnetic unit pocket to attract the magnetic microcarriers to an outer surface of the magnetic unit pocket; (l) transferring the magnetic microcarriers from the label well to the first label washing well by moving the magnetic unit pocket and magnetic unit out of the label well and into the first label washing well; A method for providing the above.

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