WO2016154113A1 - Puce graphique à barres volumétriques multiplexées pour biomarqueur hors-laboratoire et/ou quantification d'analyte, comprenant le contrôle compétitif - Google Patents

Puce graphique à barres volumétriques multiplexées pour biomarqueur hors-laboratoire et/ou quantification d'analyte, comprenant le contrôle compétitif Download PDF

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Publication number
WO2016154113A1
WO2016154113A1 PCT/US2016/023416 US2016023416W WO2016154113A1 WO 2016154113 A1 WO2016154113 A1 WO 2016154113A1 US 2016023416 W US2016023416 W US 2016023416W WO 2016154113 A1 WO2016154113 A1 WO 2016154113A1
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WO
WIPO (PCT)
Prior art keywords
recess
plate
sample
ink
row
Prior art date
Application number
PCT/US2016/023416
Other languages
English (en)
Inventor
Lidong Qin
Ying Li
Ping Wang
Xifeng Wu
Original Assignee
The Methodist Hospital
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from US14/817,258 external-priority patent/US10228369B2/en
Application filed by The Methodist Hospital filed Critical The Methodist Hospital
Priority to US15/559,968 priority Critical patent/US20180106797A1/en
Publication of WO2016154113A1 publication Critical patent/WO2016154113A1/fr

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Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
    • G01N33/54366—Apparatus specially adapted for solid-phase testing
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
    • B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip
    • B01L3/502715—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip characterised by interfacing components, e.g. fluidic, electrical, optical or mechanical interfaces
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
    • B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip
    • B01L3/502738—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip characterised by integrated valves
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/06—Fluid handling related problems
    • B01L2200/0621—Control of the sequence of chambers filled or emptied
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/06—Fluid handling related problems
    • B01L2200/0642—Filling fluids into wells by specific techniques
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
    • B01L2200/06—Fluid handling related problems
    • B01L2200/0694—Creating chemical gradients in a fluid
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0809—Geometry, shape and general structure rectangular shaped
    • B01L2300/0816—Cards, e.g. flat sample carriers usually with flow in two horizontal directions
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0861—Configuration of multiple channels and/or chambers in a single devices
    • B01L2300/0864—Configuration of multiple channels and/or chambers in a single devices comprising only one inlet and multiple receiving wells, e.g. for separation, splitting
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0861—Configuration of multiple channels and/or chambers in a single devices
    • B01L2300/0867—Multiple inlets and one sample wells, e.g. mixing, dilution
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0861—Configuration of multiple channels and/or chambers in a single devices
    • B01L2300/087—Multiple sequential chambers
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0861—Configuration of multiple channels and/or chambers in a single devices
    • B01L2300/0874—Three dimensional network
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0861—Configuration of multiple channels and/or chambers in a single devices
    • B01L2300/0883—Serpentine channels
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/04—Moving fluids with specific forces or mechanical means
    • B01L2400/0403—Moving fluids with specific forces or mechanical means specific forces
    • B01L2400/0406—Moving fluids with specific forces or mechanical means specific forces capillary forces
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/06—Valves, specific forms thereof
    • B01L2400/0622—Valves, specific forms thereof distribution valves, valves having multiple inlets and/or outlets, e.g. metering valves, multi-way valves
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2400/00—Moving or stopping fluids
    • B01L2400/06—Valves, specific forms thereof
    • B01L2400/0633—Valves, specific forms thereof with moving parts
    • B01L2400/065—Valves, specific forms thereof with moving parts sliding valves

Definitions

  • MULTIPLEXED VOLUMETRIC BAR CHART CHIP FOR POINT OF CARE BIOMARKER AND ANALYTE QUANTITATION (Attorney's Docket No. METHODIST-4), which in turn claims benefit of prior U.S. Provisional Patent Application Serial No. 61/714,676, filed 10/16/2012 by The Cincinnati Hospital Research Institute and Lidong Qin et al . for MULTIPLEXED VOLUMETRIC BAR CHART CHIP FOR POINT OF CARE BIOMARKER QUANTITATION (Attorney's Docket No. METHODIST-4 PROV) ;
  • This invention relates generally to methods and apparatus for determining the quantity of a protein and/or other biomarkers and/or analytes present in a sample, and more particularly to methods and apparatus for point of care determination of the quantity of a protein (and, preferably, the quantity of multiple biomarkers) present in a sample.
  • biomarker research has identified many helpful proteomics and genomic panels for disease diagnosis and prognosis, including cancer, infection, cardiovascular disease, diabetes, Alzheimer's disease and others. For example, a four-biomarker panel has been developed for detecting early stage ovarian cancer, and an 18-protein biomarker panel has been developed for the diagnosis of early Alzheimer's disease .
  • ELISA immunosorbent assay
  • biomarker assay where a blood sample is drawn from a patient and then processed by a relatively large, complex instrument .
  • apparatus for point of care determination of the quantity of a protein (and, preferably, the quantity of multiple proteins) present in a sample is provided.
  • apparatus for determining the quantity of a target protein and/or other types of biomarkers or analytes present in a sample, the apparatus
  • top plate comprising a plurality of recesses arranged to form a plurality of rows extending
  • a bottom plate comprising a plurality of recesses arranged to form a plurality of rows extending parallel to one another, and a plurality of channels extending perpendicularly to the plurality of rows of the bottom plate;
  • top plate and the bottom plate are assembled together so that the top plate is on top of the bottom plate and the recesses of the top plate communicate with the recesses of the bottom plate so as to form a plurality of rows;
  • top plate and the bottom plate is configured to slide relative to the other of the top plate and the bottom plate in order to form a plurality of columns, with each of the plurality of columns in communication with each of the plurality of channels.
  • a method for determining the quantity of a target protein and/or other types of biomarkers or analytes present in a sample comprising: providing apparatus comprising:
  • top plate comprising a plurality of recesses arranged to form a plurality of rows
  • a bottom plate comprising a plurality of recesses arranged to form a plurality of rows
  • top plate and the bottom plate are assembled together so that the top plate is on top of the bottom plate and the recesses of the top plate communicate with the recesses of the bottom plate so as to form a plurality of rows;
  • top plate and the bottom plate is configured to slide relative to the other of the top plate and the bottom plate in order to form a plurality of columns, with each of the plurality of columns in communication with each of the plurality of channels;
  • determining the quantity of the target protein and/or other biomarker and/or other molecular analyte present in the sample by detecting the longitudinal position of the ink contained in the plurality of channels .
  • providing apparatus comprising:
  • top plate comprising a plurality of recesses arranged to form a plurality of rows
  • a bottom plate comprising a plurality of recesses arranged to form a plurality of rows
  • top plate and the bottom plate are assembled together so that the top plate is on top of the bottom plate and the recesses of the top plate communicate with the recesses of the bottom plate so as to form a plurality of rows;
  • top plate and the bottom plate is configured to slide relative to the other of the top plate and the bottom plate in order to form a plurality of columns, with each of the plurality of columns in communication with each of the plurality of channels;
  • binding a capture agent in at least one recess forming one of the plurality of rows of the top plate introducing a sample into the at least one recess so that an analyte contained in the sample is bound to the capture agent, and binding a probe to the bound analyte; and positioning a reagent in a recess
  • determining the quantity of the analyte present in the sample by detecting the longitudinal position of the ink contained in the plurality of channels.
  • apparatus for determining the quantity of a target protein and/or other types of biomarkers or analytes present in a sample, the apparatus
  • top plate comprising at least one recess
  • bottom plate comprising at least one recess, and at least one serpentine channel communicating with the at least one recess of the bottom plate
  • At least one of the top plate and the bottom plate is configured to slide relative to the other of the top plate and the bottom plate in order to align the at least one recess of the top plate with the at least one recess of the bottom plate, so that there exists fluid communication between the at least one recess of the top plate, the at least one recess of the bottom plate, and the at least one serpentine channel communicating with the at least one recess of the bottom plate.
  • apparatus for determining the quantity of a target analyte present in a sample comprising:
  • a first plate comprising a plurality of recesses arranged to form a plurality of rows extending
  • a second plate comprising a plurality of recesses arranged to form a plurality of rows extending
  • the recesses of the first plate communicate with the recesses of the second plate so as to form a plurality of sample rows, a plurality of control rows, and an ink row disposed between the plurality of sample rows and the plurality of control rows, with the plurality of channels of the first plate being disposed between the plurality of control rows and the ink row, and the plurality of channels of the second plate being disposed between the plurality of sample rows and the ink row;
  • first plate and the second plate is configured to slide relative to the other of the first plate and the second plate in order to form a plurality of sample columns, a plurality of control columns and a plurality of ink columns, with each of the plurality of channels in the second plate being in communication with each of the plurality of sample columns and ink columns and with each of the plurality of channels in the first plate being in communication with each of the plurality of control columns and ink columns.
  • a first plate comprising a plurality of recesses arranged to form a plurality of rows
  • a second plate comprising a plurality of recesses arranged to form a plurality of rows
  • first plate and the second plate are assembled together so that the first plate is positioned against the second plate and the recesses of the first plate communicate with the recesses of the second plate so as to form a plurality of sample rows, a plurality of control rows, and an ink row disposed between the plurality of sample rows and the plurality of control rows, with the plurality of channels of the first plate being disposed between the plurality of control rows and the ink row, and the plurality of channels of the second plate being disposed between the plurality of sample rows and the ink row; and
  • first plate and the second plate is configured to slide relative to the other of the first plate and the second plate in order to form a plurality of sample columns, a plurality of control columns and a plurality of ink columns, with each of the plurality of channels in the second plate being in communication with each of the plurality of sample columns and ink columns and with each of the plurality of channels in the first plate being in communication with each of the plurality of control columns and ink columns;
  • determining the quantity of the target analyte present in the sample by detecting the disposition of the ink contained in the plurality of channels in the second plate and the plurality of channels in the first plate.
  • apparatus for determining the quantity of a target analyte present in a sample comprising:
  • control recess a control recess, a sample recess, an ink recess disposed between the control recess and the sample recess, and a channel fluidically connecting the control recess, the ink recess and the sample recess; a known concentration of a reactant being
  • an analyte-specific antibody being bound in the sample recess, and ink being disposed in the ink recess;
  • providing apparatus comprising:
  • control recess a control recess, a sample recess, an ink recess disposed between the control recess and the sample recess, and a channel fluidically connecting the control recess, the ink recess and the sample recess ;
  • determining the quantity of the target analyte present in the sample by detecting the disposition of the ink in the channel.
  • apparatus for determining the quantity of a target analyte present in a sample comprising:
  • a first plate comprising a first surface having a first recess containing an analyte-specific antibody and a second recess for containing ink, the first recess being spaced from the second recess;
  • a second plate comprising a second surface having a third recess for containing a reagent, a fourth elongated recess and a fifth recess, the fifth recess being disposed between, and spaced from, the third recess and the fourth elongated recess;
  • first plate and the second plate are assembled together so that the first surface of the first plate faces the second surface of the second plate ;
  • first plate and the second plate are reconfigurable between (i) a first state in which the first recess is fluidically isolated from the third recess and the fifth recess and the second recess is fluidically isolated from the fourth elongated recess and the fifth recess, and (ii) a second state in which the first recess is fluidically connected to the third recess and the fifth recess and the second recess is fluidically connected to the fourth elongated recess and the fifth recess.
  • providing apparatus comprising:
  • a first plate comprising a first surface having a first recess containing an analyte-specific antibody and a second recess for containing ink, the first recess being spaced from the second recess;
  • a second plate comprising a second surface having a third recess for containing a reagent, a fourth elongated recess and a fifth recess, the fifth recess being disposed between, and spaced from, the third recess and the fourth elongated recess;
  • first plate and the second plate are assembled together so that the first surface of the first plate faces the second surface of the second plate ;
  • first plate and the second plate are reconfigurable between (i) a first state in which the first recess is fluidically isolated from the third recess and the fifth recess and the second recess is fluidically isolated from the fourth elongated recess and the fifth recess, and (ii) a second state in which the first recess is fluidically connected to the third recess and the fifth recess and the second recess is fluidically connected to the fourth elongated recess and the fifth recess;
  • Fig. 1 which shows a typical prior art approach for a protein-based biomarker assay, where a blood sample is drawn from a patient and then processed by a relatively large, complex instrument;
  • Fig. 2 shows the novel multiplexed volumetric bar chart chip of the present invention
  • Fig. 3 shows the novel multiplexed volumetric bar chart chip of Fig. 2 and a barcode scanner which can be used to read the multiplexed volumetric bar chart chip;
  • Figs. 4-8 illustrate further details of the novel multiplexed volumetric bar chart chip of the present invention
  • Fig. 9 is a schematic drawing of an etching process which can be utilized to form recesses and channels in the top plate and the bottom plate of multiplexed volumetric bar chart chip;
  • Fig. 10 is a schematic drawing of the assembly and operation of the multiplexed volumetric bar chart chip of the present invention.
  • Figs. 11 and 12 are schematic drawings
  • Fig. 13 shows the multiplexed volumetric bar chart chip of the present invention prior to the oblique sliding of the top plate relative to the bottom plate;
  • Figs. 14-16 show the test results obtained in accordance with the present invention for various samp1es ;
  • Figs. 17-20 show specific steps which are
  • Figs. 21-32 are a schematic series of views illustrating the assembly and operation of the
  • Figs. 33-45 are a schematic series of views showing how, over time, the ink in various bar
  • channels advance in the multiplexed volumetric bar chart chip according to the quantity of target
  • Fig. 46 illustrates specific steps which are performed in accordance with a DNA assay scheme and oxygen generation mechanism
  • Fig. 47 shows an alternative embodiment of the novel multiplexed volumetric bar chart chip of the present invention.
  • Fig. 48 shows images of hydrogen peroxide
  • Figs. 49 and 50 show an alternative embodiment of the novel multiplexed volumetric bar chart chip of the present invention
  • Fig. 51 shows an alternative embodiment of the novel multiplexed volumetric bar chart chip of the present invention
  • Fig. 52 shows an alternative embodiment of the novel multiplexed bar chart chip of the present invention, wherein platinum nanoparticles are utilized in the place of catalase;
  • Fig. 53 shows an alternative embodiment of the novel multiplexed bar chart chip of the present invention, wherein the channel (s) are arranged in a serpentine configuration;
  • Fig. 54 shows an alternative embodiment of the novel multiplexed bar chart chip of the present invention, wherein the channels are arranged in straight and V-shaped configurations;
  • Figs. 54A and 54B are schematic views showing another alternative form of multiplexed volumetric bar chart chip formed in accordance with the present invention.
  • Fig. 54C is a schematic view showing further details of the top plate of the multiplexed volumetric bar chart chip of Figs. 54A and 54B;
  • Fig. 54D is a schematic view showing further details of the bottom plate of the multiplexed
  • Fig. 54E is a schematic view of the assembly of the top plate and the bottom plate of the multiplexed volumetric bar chart chip of Figs. 54A and 54B;
  • Fig. 54F is a schematic view showing a modified form of the multiplexed volumetric bar chart chip of Figs. 54A and 54B;
  • Figs. 54G-54I are schematic views illustrating the principles of various assays which may be used with the multiplexed volumetric bar chart chip of the present invention.
  • Figs. 54J-54M are schematic views illustrating use of the multiplexed volumetric bar chart chip of Figs. 54A and 54B;
  • Fig. 54N is a table listing exemplary threshold cutoff values for various analytes which may be assayed using the multiplexed volumetric bar chart chip of Figs. 54A and 54B;
  • Fig. 540 is a schematic view showing still another alternative form of multiplexed volumetric bar chart chip formed in accordance with the present invention.
  • Fig. 54P is a schematic view showing further details of the top plate of the multiplexed volumetric bar chart chip of Fig. 540;
  • Fig. 54Q is a schematic view showing further details of the bottom plate of the multiplexed
  • Figs. 54R-54T are schematic views of the assembly of the top plate and the bottom plate of the
  • FIG. 540 Figs. 54U-54W are schematic views illustrating use of the multiplexed volumetric bar chart chip of Fig. 540;
  • Fig. 55 shows an alternative embodiment of the novel multiplexed bar chart chip of the present invention, wherein the novel multiplexed bar chart chip is configured for a hepatocellular carcinoma risk assessment assay;
  • Fig. 56 shows an alternative embodiment of the novel multiplexed bar chart chip of the present invention, wherein the novel multiplexed bar chart chip is configured for a breast cancer risk/diagnosis assay;
  • Fig. 57 shows an alternative embodiment of the novel multiplexed bar chart chip of the present invention, wherein the novel multiplexed bar chart chip is configured for a sepsis assessment assay;
  • Fig. 58 shows an alternative embodiment of the novel multiplexed bar chart chip of the present invention, wherein the novel multiplexed bar chart chip is configured for a drug abuse asssessment assay;
  • Fig. 59 shows an alternative embodiment of the novel multiplexed bar chart chip of the present invention, wherein the novel multiplexed bar chart chip is configured for assay of several important physiological biomarkers;
  • Fig. 60 shows an alternative embodiment of the novel multiplexed bar chart chip of the present invention, wherein the novel multiplexed bar chart chip is configured for an assay of DNA, RNA, and/or micro-RNA targets.
  • the present invention provides a new method and apparatus for point of care determination of the quantity of a protein (and, preferably, the quantity of multiple proteins) present in a sample.
  • Multiplexed volumetric bar chart chip 5 is configured to simultaneously determine the quantity of multiple proteins which may be present in a sample, with the quantity of each protein which is present in the sample being indicated in a particular one of a plurality of bar channels 10.
  • 6 10, 30, and 50-plexed, or more than 50-plexed, channels may be incorporated into
  • Bar channels 10 may be straight (as shown in Fig. 2) or curved (e.g., serpentine, circular, z-shaped) or formed in any other configuration which provides a series of channels having a length. As a result of this
  • the review of a particular bar channel 10 will indicate the quantity of a particular protein which may be present in the sample and, significantly, the collective array of the plurality of bar channels 10 will simultaneously indicate, in bar chart form, the quantities of multiple proteins which may be present in the sample, whereby to provide multi- protein quantity measurements and hence a more
  • the multi-protein measurements presented in bar chart form by multiplexed volumetric bar chart chip 5 may then be read with a smart-phone or barcode scanner 15, whereby to automate the data collection process.
  • multiplexed volumetric bar chart chip 5 comprises two plates, a transparent top plate 20 and a bottom plate 25 (which may or may not be transparent) .
  • Top plate 20 (Figs. 5 and 6) has a plurality of recesses 30 formed on its bottom surface, with
  • each of the recesses 30 extending at a 45 degree angle relative to the axis of a given row 35, and with a recess 30 in one row 35 being aligned with an offset recess 30 in an adjacent row 35.
  • An inlet 40 is connected to a far side recess
  • An inlet 50 is connected to a far side recess 30 on the penultimate row 35B, and an outlet 55 is formed adjacent to the opposite far side recess 30 on the same penultimate row 35B.
  • the antepenultimate row 35C lacks both an inlet and an outlet.
  • An inlet 60 is connected to a far side recess
  • recesses 30, inlets 40, 50, 60, and outlets 45, 55, 65 are all formed in the bottom surface of top plate 20 using a conventional etching process of the sort well known in the etching arts.
  • recesses 30, inlets 40, 50, 60 and outlets 45, 55, 65 are etched in the bottom surface of a glass plate.
  • recesses 30, inlets 40, 50, 60 and outlets 45, 55, 65 may be formed in a silicon plate, a plastic plate, a ceramic plate, a quartz plate, a metal oxide plate or other appropriate substrate material.
  • Bottom plate 25 has a plurality of recesses 70 formed on its top surface, with recesses 70 being arranged in a plurality of rows 75 (i.e., 75A, 75B, 75C, etc.), with each of the recesses 70 extending at a 45 degree angle relative to the axis of a given row
  • the plurality of bar channels 10 are formed on the top surface of bottom plate 25, with each of the bar channels 10 being connected to a recess 70 in the ante-ante- antepenultimate row 75E (see Fig. 8), and with each of the bar channels 10 extending parallel to one another and perpendicular to the axis of rows 75.
  • recesses 70, outlets 80, 85, 90, and bar channels 10 are all formed in the top surface of bottom plate 25 using a conventional etching process of the sort well known in the etching arts.
  • recesses 70, outlets 80, 85, 90, and bar channels 10 are etched in the top surface of a glass plate.
  • recesses 70, outlets 80, 85, 90, and bar channels 10 may be formed in a silicon plate, a plastic plate, a ceramic plate, a quartz plate, a metal oxide plate or other appropriate substrate material.
  • top plate 20 is
  • top plate 20 is assembled on top of bottom plate 25 so that recesses 30 in top plate 20 communicate with recesses 70 in bottom plate 25. More particularly, when top plate 20 is assembled on top of bottom plate 25 in this manner, recesses 30 in top plate 20 will cooperate with recesses 70 in bottom plate 25 so as to initially form a plurality of continuous rows 95 (i.e., 95A, 95B,
  • the antepenultimate row 95C lacks both an inlet and an outlet.
  • each column 100 being in fluid communication with one of the aforementioned bar columns 10.
  • multiplexed volumetric bar chart chip 5 can be used to simultaneously determine the quantity of multiple proteins present in a sample, with the quantity of each specific protein being indicated in a particular one of the plurality of bar channels 10.
  • row 75B will contain a series of different protein-specific antibodies, with a different protein-specific antibody being located in each recess 30 of the row 75B.
  • Red ink (or some other colored material which is readily discernible through top plate 25 and against bottom plate 20) is introduced into inlet 60 of multiplexed volumetric bar chart chip 5, whereby to fill the ante-antepenultimate row 75D of multiplexed volumetric bar chart chip 5 with red ink.
  • Antepenultimate row 75C is intentionally left blank to serve as an air spacer, thereby avoiding direct contact between a sample and the red ink.
  • the sample is introduced into inlet 50 of multiplexed volumetric bar chart chip 5 so that the sample fills the penultimate row 75B.
  • This action causes the sample to mix with the different protein- specific antibodies which are bonded to bottom plate 20 in the recesses 30, so that the target proteins bind to the appropriate protein-specific antibodies in the recesses 30.
  • each target protein binds to only one protein-specific antibody, and such binding takes place in only one of the recesses 30 in the penultimate row 75B.
  • the penultimate row 75B is flushed so as to remove any materials which are not bound to a protein-specific antibody.
  • catalase is introduced into inlet 50 of multiplexed volumetric bar chart chip 5 so as to fill the penultimate row 75B.
  • This action causes the catalase to bind to the target proteins which are themselves bound to the protein-specific antibodies in the recesses 30.
  • the catalase is a mixture of all the catalase detecting probes required for binding to the target proteins (e.g., silica nanoparticles conjugated with detecting antibodies and catalase molecules) .
  • excess catalase is rinsed from the penultimate row 75B.
  • top plate 25 is slid obliquely relative to bottom plate 20, causing rows 75 (i.e.,
  • each recess 30 (containing the protein-specific antibodies and any target proteins bound thereto and any catalase bound thereto) previously located in penultimate row 75B becomes incorporated as a section of a specific column 100 (i.e., 100A, 100B, lOOC, etc.) .
  • this row-to-column transformation occurs, each recess 30 (containing the protein-specific antibodies and any target proteins bound thereto and any catalase bound thereto) previously located in penultimate row 75B becomes incorporated as a section of a specific column 100 (i.e., 100A, 100B, lOOC, etc.) .
  • the hydrogen peroxide contained in row 75A is permitted to advance up each of the columns 100 and thereby mix with any catalase bound to the target proteins (which are themselves bound to the protein-specific antibodies), the mixing of which causes a reaction which releases oxygen gas.
  • the oxygen gas is produced in proportion to the quantity of catalase present in a given column (and hence in proportion to the quantity of target proteins which are present in a given column) .
  • the quantity of oxygen gas produced in a given column 100 is
  • each of the columns 100 contains a different target protein (by virtue of the fact that each of the columns 100 contains a different protein-specific antibody) .
  • the oxygen gas produced by the reaction accumulates within the limited volume of columns 100 and causes an increase in pressure, which propels the red ink contained in columns 100 into and along bar columns 10, with the ink advancing a distance along bar columns 10 which is proportional to the quantity of oxygen gas produced in that column, which is in turn proportional to the quantity of the target proteins which are bound to the protein-specific antibodies disposed in the recesses associated with that column.
  • multiplexed volumetric bar chart chip 5 can be used to simultaneously determine the quantity of multiple proteins present in a sample, with the quantity of each protein being indicated in a particular one of a plurality of bar channels 10. See, for example, Figs.
  • Fig. 13-16 shows multiplexed volumetric bar chart chip 5 prior to the oblique sliding of top plate 25 relative to bottom plate 20, and Figs. 14-16 show the test results for various samples.
  • Figs. 17-20 show specific steps in the foregoing process. Specifically, Fig. 17 shows a protein- specific antibody being bound in a recess 30 of bottom plate 20; Fig. 18 shows a sample being loaded into a recess 30 of bottom plate 20, whereby to bind a target protein to a protein-specific antibody; Fig. 19 shows catalase being loaded into a recess 30 so as to bind catalase to a target protein (which is itself bound to a protein-specific antibody); and Fig. 20 shows hydrogen peroxide being loaded into a recess 30, whereby to release oxygen gas in proportion to the quantity of target protein present in a recess 30.
  • the same protein-specific antibody can be bound in multiple recesses 30 of penultimate row 35B of bottom plate 20, whereby to provide
  • Figs. 21-32 are a schematic series of views showing the assembly and operation of the multiplexed volumetric bar chart chip in one preferred form of the present invention.
  • Figs. 33-45 are a schematic series of views showing how, over time, the ink in a given bar channel advances a distance along that bar channel which is proportional to the quantity of the target protein which are bound to the protein-specific antibody disposed in the recess associated with that bar channel, whereby to indicate, in multiplexed
  • novel method and apparatus of the present invention provides instant and visual quantitation of target biomarkers or other molecular analytes and provides a visualized bar chart without the use of instruments, data processing or graphic plotting.
  • the novel method and apparatus of the present invention can be easily used as a point of care determination of the quantity of a protein (and, preferably, the quantity of multiple proteins) present in a sample. More particularly, the novel method and apparatus of the present invention can be used as a point of care determination of the quantity of protein, nucleic acid, peptide, sugar, organic compounds, polymer, metal ions, and/or other molecular analytes, as well as the quantity of bacteria, cells, and/or particles.
  • gas is generated by the reaction of an ELISA probe with a reagent, and specifically, gas is generated by the reaction of the
  • ELISA probe i.e., the protein-specific antibody which is bound to the target protein which is bound to the catalase
  • hydrogen peroxide hydrogen peroxide
  • the multiplexed volumetric bar chart chip is based on a sandwich assay.
  • a capture antibody binds to an analyte and a detecting antibody conjugated with a catalase probe indicates the amount.
  • the sandwich scheme is made up of capture antibody/analyte/detecting antibody conjugated with a catalase probe.
  • This type of sandwich scheme could also be extended to nucleic acid hybridization, where the sandwich is capture DNA strand/target strand/detecting
  • DNA strand i.e., the target strand has a first half complimentary to the capture DNA strand and a second half complimentary to the detecting DNA strand
  • Fig. 46 shows specific steps that are performed in accordance with a DNA assay scheme and oxygen generation
  • this type of sandwich scheme could also be extended to hydrogen bonding, electrostatic reaction or interaction, or covalent bonding, where the target analyte is captured by a surface with a coating that can adhere the analyte by either hydrogen bonding, electrostatic reaction or interaction or the formation of a covalent bond.
  • the readout of the adhered or bonded analyte can then be detected by the detecting antibody with a catalase probe.
  • sandwich of these types are surfaces (with adhesion forces of hydrogen bonding, electrostatic interaction or covalent bonding) /analyte/probe of detecting antibody with catalase.
  • a novel multiplexed volumetric bar chart chip 200 is provided which may be used in accordance with the present invention to determine the quantity of a target protein or other types of biomarkers or other analytes, wherein the signal for determining the quantity of the target protein or other types of biomarkers or other analytes is amplified.
  • multiplexed volumetric bar chart chip 200 comprises two glass plates, a transparent top plate 220 and a bottom plate 225
  • Top plate 220 and bottom plate 225 are similar to top plate 20 and bottom plate 25 discussed above, except that the plurality of rows are arranged on the multiplexed volumetric bar chart chip 200 so that the recesses in the rows are filled with the ELISA
  • reagents i.e., the protein-specific antibody, with the sample and catalase bound thereto
  • hydrogen peroxide platinum film, hydrogen peroxide, platinum film, hydrogen peroxide, platinum film and ink.
  • oxygen is generated, with that oxygen being proportional to the quantity of the target antibody present in the sample.
  • the oxygen generated by the ELISA reaction in turn drives a quantity of unreacted hydrogen peroxide (that is proportional to the
  • multiplexed volumetric bar chart chip 200 exhibits a higher sensitivity than the multiplexed volumetric bar chart chip 5 discussed above. See, for example, Fig. 48, which shows images of hydrogen peroxide solution being pushed into successive
  • a novel multiplexed volumetric bar chart chip 300 is provided.
  • Multiplexed volumetric bar chart chip 300 is similar to multiplexed volumetric bar chart chip 5 discussed above, except that multiplexed volumetric bar chart chip 300 is manufactured so as to reduce the reagent loading and washing steps required for a user.
  • the ELISA reagents i.e., the washing buffer, catalase probe and washing buffer
  • the multiplexed volumetric bar chart chip can be pre-loaded in the multiplexed volumetric bar chart chip during the manufacturing stage (e.g., at the locations shown in Fig. 49) .
  • the sample is positioned in the multiplexed volumetric bar chart chip (e.g., at the location shown in Fig. 49) .
  • the multiplexed volumetric bar chart chip is slid vertically so that the sample, washing buffer, catalase probe and washing buffer are sequentially passed through the ELISA reagent row of the chip, whereby to prepare the ELISA row of the chip in a single action. Subsequently, the multiplexed
  • volumetric bar chart chip can be slid in the oblique direction so as to activate the oxygen reaction and generate the desired results.
  • the user will only need to load the sample into the chip and then slide the chip obliquely so as to activate the assay
  • Fig. 51 shows another form of the present
  • the multiplexed volumetric bar chart chip is configured to load the ELISA row of the chip through a horizontal motion.
  • gas is generated by the reaction of an ELISA probe with a reagent, and specifically, gas is generated by the reaction of the ELISA probe (i.e., the protein-specific antibody which is bound to the target protein which is bound to the catalase) with hydrogen peroxide.
  • the ELISA probe i.e., the protein-specific antibody which is bound to the target protein which is bound to the catalase
  • hydrogen peroxide hydrogen peroxide
  • platinum nanoparticles may be utilized in the place of catalase.
  • a sample e.g., blood, urine, etc.
  • inlet 50 of multiplexed volumetric bar chart chip 5 so that the sample fills the penultimate row 75B, causing the sample to mix with the different protein-specific antibodies which are bonded to bottom plate 20 in recesses 20, so that the target proteins bind to the appropriate protein-specific antibodies in the recesses 30.
  • Row 75B is then flushed (e.g., with a buffer solution) so as to remove any materials which are not bound to a protein-specific antibody.
  • Platinum nanoparticles conjugated with detection antibodies are then added into inlet 50 so as to fill penultimate row 75B. This action causes the platinum nanoparticles to bind to the target proteins which are themselves bound to the protein-specific antibodies in the recesses 30. The platinum nanoparticles are then rinsed from penultimate row 75B, leaving behind only those platinum nanoparticles which are bound to target proteins via their detection antibodies.
  • top plate 25 is slid obliquely relative to bottom plate 20, rows 75 are transformed into columns 100, and hydrogen peroxide contained in row 75A is permitted to advance up each of the columns
  • Platinum nanoparticles exhibit several properties which can make them advantageous.
  • catalase reacts with hydrogen peroxide for up to about 2 minutes, whereas platinum nanoparticles have no such limitation.
  • platinum nanoparticles can provide higher sensitivity and longer stability than catalase.
  • bar channels 10 are generally discussed in the context of a bar chart, where a plurality of straight bar channels 10 are arranged in parallel so as to provide a series of discrete channels.
  • bar channels 10 may be curved (e.g., serpentine, circular, z-shaped) or formed in other configuration which provides a series of channels having a length.
  • bar channels 10 may comprise a single
  • serpentine pathway 410 (see Fig. 53) .
  • Providing a single serpentine pathway may be advantageous in situations where the target protein is present in a high concentration and quantitation is desired (e.g., as may be desired in a pregnancy test) .
  • channels 10 may vary along the length of the channels.
  • channels 10 may comprise a V-shape, where the distal end (i.e., the terminal portion) of channel 10 is of greater width than the width of the channel at its proximal end.
  • the depth of channel 10 may also be varied along the length of the channel (e.g., to provide a deeper channel 10 toward the distal end of the channel) .
  • the volume of the interior of the channel may be varied, whereby to provide additional time/space for the advancement of the red ink during a reaction.
  • construction may be advantageous to obtain a higher sensitivity and a larger dynamic range for a desired assay .
  • a novel multiplexed volumetric bar chart chip Utilizing "Competitive" Control
  • a threshold quantity of a target protein or other types of biomarkers or other
  • a reaction with a control is used to generate a gas that acts in direct competition with a gas that is generated by a reaction with a sample, whereby to provide a multiplexed volumetric bar chart chip which exhibits a clear positive or negative indication of the presence of an analyte (i.e., the ink moves from a central location in a column on the multiplexed volumetric bar chart chip into either the "positive" side of the multiplexed volumetric bar chart chip or the "negative" side of the multiplexed volumetric bar chart chip) .
  • invention also allows for the setting of a
  • analyte i.e., the concentration of the control can be selected so as to reduce false positives when the concentration of the analyte is extremely low.
  • this form of the invention minimizes the influence of environmental conditions (e.g., temperature, humidity, pH, ionic strength, etc.) on the assay, and eliminates the need for calibration of the multiplexed volumetric bar chart chip, by placing the control and the sample in direct "competition" with one another. Put another way, since the control and the sample are subject to the same environmental conditions, the effects of those environmental conditions are effectively
  • a novel multiplexed volumetric bar chart chip 500 is provided.
  • Multiplexed volumetric bar chart chip 500 is
  • bar channels 505 may be incorporated into multiplexed volumetric bar chart chip 500 (see, for example, Fig. 54F which shows a 20- plexed volumetric bar chart chip 500) .
  • Bar channels may be incorporated into multiplexed volumetric bar chart chip 500 (see, for example, Fig. 54F which shows a 20- plexed volumetric bar chart chip 500) .
  • 505 may be straight or curved (e.g., serpentine, circular, z-shaped) or formed in any other
  • channels preferably have the same configuration and orientation.
  • the review of a particular bar channel 505 will indicate (i) whether an analyte is present in a sample at a concentration above a predetermined threshold value, and (ii) the quantity of a particular analyte which may be present in the sample and, significantly, the collective array of the plurality of bar channels 505 will simultaneously indicate, in bar chart form, the presence of and quantities of multiple analytes which may be present in the sample, whereby to provide multi-analyte quantity measurements and hence a more comprehensive diagnostic result.
  • Figs. 54C, 54D and 54E Looking now at Figs. 54C, 54D and 54E,
  • multiplexed volumetric bar chart chip 500 comprises two plates, a transparent top plate 510 and a bottom plate 515 (which may or may not be transparent) .
  • top plate 510 (Fig. 54C) has a plurality of recesses 520 formed on its bottom
  • each of the recesses 520 extends at a 45 degree angle relative to the axis of a given sample row 525 or control row 535, with each recess 520 in one row 525, 535 being aligned with an offset recess 520 in an adjacent row 525, 535.
  • An inlet 545 is connected to a far side recess 520 on the ultimate sample row 525A, and an outlet 550 is formed adjacent to the opposite far side recess 520 on the same ultimate sample row 525A.
  • An inlet 555 is connected to a far side recess 520 on the penultimate sample row 525B, and an outlet 560 is formed adjacent to the opposite far side recess 520 on the same penultimate sample row 525B.
  • the antepenultimate sample row 525C lacks both an inlet and an outlet.
  • An inlet 565 is connected to a far side recess 520 on the ultimate control row 535A, and an outlet 570 is formed adjacent to the opposite far side recess 520 on the same ultimate control row 535A.
  • An inlet 575 is connected to a far side recess 520 on the penultimate control row 535B, and an outlet 580 is formed adjacent to the opposite far side recess 520 on the same penultimate control row
  • antepenultimate control row 535C lacks both an inlet and an outlet.
  • a plurality of recesses 520 are also formed on the bottom surface of top plate 510 intermediate sample portion 530 and control portion 540, whereby to form an ink row 585.
  • An inlet 590 is connected to a far side recess 520 on ink row 585, and an outlet 595 is formed adjacent to the opposite far side recess 520 on ink row 585.
  • a plurality of bar channels 600 are formed on the bottom surface of top plate 510, with each of the bar channels 600 being connected to a recess 520 in the antepenultimate row 535C of control portion 540, and with each of the bar channels 600 extending from antepenultimate row 535C toward ink row 585, parallel to one another and perpendicular to the axis of rows 525, 535.
  • recesses 520, inlets 545, 555, 565, 575, 590, outlets 550, 560, 570, 580, 595, and bar channels 600 are all formed in the bottom surface of top plate 510 using a
  • recesses 520, inlets 545, 555, 565, 575, 590, outlets 550, 560, 570, 580, 595 and bar channels 600 are etched in the bottom surface of a glass plate.
  • recesses 520, inlets 545, 555, 565, 575, 590, outlets 550, 560, 570, 580, 595 and bar channels 600 may be formed in a silicon plate, a plastic plate, a ceramic plate, a quartz plate, a metal oxide plate or other appropriate substrate material.
  • Bottom plate 515 (Fig. 54D) has a plurality of recesses 605 formed on its top surface, with recesses 605 being arranged in a plurality of sample rows 610 (i.e., 610A, 610B and 610C) arrayed along the lower
  • each of the recesses 605 extends at a 45 degree angle relative to the axis of a given sample row 610 or control row 620, with each recess 605 in one row 610, 620 being aligned with an offset recess 605 in an adjacent row 610, 620.
  • An outlet 630 is connected to a far side recess
  • An outlet 635 is connected to a far side recess 605 on the penultimate sample row 610B.
  • the antepenultimate sample row 610C lacks both an inlet and an outlet.
  • An outlet 640 is connected to a far side recess 605 on the ultimate control row 620A.
  • An outlet 645 is connected to a far side recess 605 on the penultimate control row 620B.
  • the antepenultimate control row 620C lacks an both an inlet and an outlet.
  • a plurality of recesses 605 are also formed on the top surface of bottom plate 515 intermediate sample portion 615 and control portion 625, whereby to form an ink row 650.
  • An outlet 655 is connected to a far side recess 605 on the ink row 650.
  • a plurality of bar channels 660 are formed on the top surface of bottom plate 515, with each of the bar channels 660 being connected to a recess 605 in the antepenultimate row 610C of sample portion 615, and with each of the bar channels 660 extending from antepenultimate row 610C toward ink row 650, parallel to one another and perpendicular to the axis of rows 610, 620.
  • outlets 630, 635, 640, 645, 655 and bar channels 660 are all formed in the top surface of bottom plate 515 using a conventional etching process of the sort well known in the etching arts.
  • recesses 605, outlets 630, 635, 640, 645, 655 and bar channels are formed in the top surface of bottom plate 515 using a conventional etching process of the sort well known in the etching arts.
  • 660 are etched in the top surface of a glass plate.
  • recesses 605, outlets 630, 635, 640, 645, 655 and bar channels 660 may be formed in a silicon plate, a plastic plate, a ceramic plate, a quartz plate, a metal oxide plate or other appropriate substrate material. Assembly Of Multiplexed Volumetric Bar Chart Chip 500
  • top plate 510 is assembled on top of bottom plate 515 so that recesses 520 in top plate 510 communicate with recesses 605 in bottom plate 515. More particularly, when top plate 510 is assembled on top of bottom plate 515 in this manner, recesses 520 in top plate 510 will cooperate with recesses 605 in bottom plate 515 so as to
  • sample rows 665 i.e., 665A, 665B, and 665C
  • sample portion 670 of multiplexed volumetric bar chart chip 500
  • control rows 675 i.e., 675A, 675B and 675C arrayed along the upper (i.e., "control") portion 680 of multiplexed volumetric bar chart chip 500, and a continuous ink row 685 in multiplexed volumetric bar chart chip 500.
  • inlet 545 of ultimate sample row 665A is connected with the outlet 550 of ultimate sample row 665A
  • inlet 555 of the penultimate sample row 665B is connected with the outlet 560 of penultimate sample row 665B
  • inlet 565 of ultimate control row 675A is connected with outlet 570 of ultimate control row 675A
  • inlet 575 of penultimate control row 675B is connected with outlet 580 of penultimate control row 675B
  • inlet 590 of ink row 685 is connected with outlet 595 of ink row 685.
  • Antepenultimate sample row 665C and antepenultimate control row 675C lack both an inlet and an outlet.
  • top plate 510 dispositions of recesses 520 in top plate 510 and recesses 605 in bottom plate 515, an oblique slide of top plate 510 relative to bottom plate 515 disrupts the aforementioned rows 665, 675, 685 and causes them to transform into a plurality of continuous columns (i.e., bar channels 505), with each bar channel 505 being in fluid communication with the aforementioned bar channels 600, 660. See Fig. 54B.
  • multiplexed volumetric bar chart chip 500 can be used to simultaneously determine whether a predetermined threshold quantity of a target protein or other types of biomarkers or other analytes is present in a sample, and the quantity of multiple analytes present in a sample, with the quantity of each specific analyte being indicated in a particular one of the plurality of bar channels 505. More particularly, and referring now to Figs. 54G, 54H and 541, and as will hereinafter be discussed in further detail below, during manufacture of
  • an analyte-specific antibody is bonded in a recess 520 of the penultimate sample row 525B of top plate 510 (and/or, if desired, in a recess 605 of the penultimate sample row 610B of bottom plate 515) . If desired, an
  • analyte-specific antibody may also be bonded in a recess 520 of the penultimate control row 535B of top plate 510 (and/or, if desired, in a recess 605 of the penultimate control row 620B of bottom plate 515) .
  • a different analyte-specific antibody is bonded in each recess 520 of top plate 510.
  • continuous penultimate sample row 665B will contain a series of different analyte-specific antibodies, with a different analyte-specific antibody being located in each recess 520 of the continuous penultimate sample row 665B of top plate 510.
  • luminol may be added to the hydrogen peroxide before the hydrogen peroxide is introduced into inlets 545, 565.
  • Red ink (or some other colored material which is readily discernible through top plate 510 and against bottom plate 515) is introduced into inlet 590 of multiplexed volumetric bar chart chip 500, whereby to fill continuous ink row 685 of multiplexed volumetric bar chart chip 500 with red ink.
  • Continuous antepenultimate sample row 665C and continuous antepenultimate control row 675C are intentionally left empty to serve as an air spacer.
  • the target analyte i.e., the analyte being tested for
  • the target analyte will bind the analyte-specific antibody which is bound to a recess 520 of continuous
  • a probe e.g., an
  • the sample is introduced into inlet 555 (Fig. 54E) of multiplexed volumetric bar chart chip 500, so that the sample fills continuous penultimate sample row 665B. After a period of time, the
  • continuous penultimate sample row 665B is washed, leaving the target analyte (s) (where present) bound to the analyte-specific antibodies which are, in turn, bound to a recess 520 of continuous penultimate sample row 665B.
  • the amount of the target analyte retained in continuous penultimate sample row 665B is proportional to the amount of the target analyte present in the sample.
  • a conjugate comprising horseradish peroxidase (HRP) bound to a detection antibody (which detection antibody is selected becaue it will bind the target analyte) is prepared.
  • the HRP-detection antibody conjugate is introduced into inlet 555 of multiplexed bar chart chip 500, so that the HRP-detection antibody conjugate fills continuous penultimate sample row
  • the HRP- detection antibody conjugate will bind the target analyte.
  • Continuous penultimate sample row 665B is then washed, leaving only the HRP-detection antibody conjugate where it is bound to the target analyte.
  • the amount of HRP present and bound in a recess 520 is proportional to the amount of target analyte bound to multiplexed volumetric bar chart chip 500, and hence, proportional to the amount of target analyte present in the sample.
  • the target analyte i.e., the analyte being tested for
  • the target analyte will bind to the analyte- specific antibody which is bound to a recess 520 of continuous penultimate sample row 665B
  • an HRP- drug derivative conjugate will bind to the analyte- specific antibody which is bound to a recess 520 of continuous penultimate sample row 665B only where the analyte-specific antibody has not already bound the target analyte, as will hereinafter be discussed in greater detail.
  • the sample is introduced into inlet 555 (Fig. 54E) of multiplexed volumetric bar chart chip 500 so that the sample fills continuous penultimate sample row 665A.
  • inlet 555 Fig. 54E
  • continuous penultimate sample row 665A is washed, leaving the target analyte (s) (where present) bound to the
  • analyte-specific antibodies which are, in turn, bound to a recess 520 of continuous penultimate sample row 665B.
  • the amount of the target analyte retained in sample row 665B is proportional to the amount of target analyte present in the sample.
  • a conjugate comprising horseradish peroxidate (HRP) bound to a drug derivative (which drug
  • the HRP-drug derivative conjugate is introduced into inlet 555 of multiplexed bar chart chip 500.
  • the HRP-drug derivative conjugate will only bind the analyte-specific antibodies where the target analyte is absent (i.e., where the target analyte has not bound to the analyte-specific antibodies) .
  • the amount of HRP present and bound in recess (es) 520 of continuous penultimate sample row 665B is inversely proportional to the amount of target analyte present in the sample.
  • hydrogen peroxide from continuous ultimate sample row 665A is thereafter introduced into continuous penultimate sample row 665B
  • continuous penultimate control row 675B (Fig. 54E) will contain a predetermined amount of HRP. More particularly, a solution containing HRP is prepared, with the concentration of HRP being selected so as to reflect the target "threshold" for detecting the target analyte, as will hereinafter be discussed in greater detail.
  • the HRP solution is introduced into inlet 575 of multiplexed volumetric bar chart chip 500, whereby to fill continuous penultimate control row 675B.
  • the reaction between the hydrogen peroxide and the HRP will generate nitrogen gas in an amount proportional to the amount of HRP present (i.e., proportional to the
  • top plate 510 is slid obliquely relative to bottom plate 515, causing continuous sample rows 665, continuous control rows 670 and continuous ink row 675 to be disrupted and transformed into continuous bar channels 505 (i.e., 505A, 505B, 505C, etc.), such as shown in Fig. 54B.
  • continuous bar channels 505 i.e., 505A, 505B, 505C, etc.
  • each recess 520 (containing the analyte-specific antibodies and any target analytes bound thereto and any HRP bound thereto) previously located in continuous penultimate sample row 665B or continuous penultimate control row 675B becomes incorporated as a section of a specific bar channel 505 (i.e., 505A, 505B, 505C, etc.).
  • the hydrogen peroxide contained in continuous ultimate sample row 665A and continuous ultimate control row 675A is permitted to advance up each of the bar channels 505 and thereby mix with any HRP bound to the analyte- specific antibodies, the mixing of which causes a reaction which releases nitrogen gas.
  • the nitrogen gas is produced in proportion to the quantity of HRP present in a given penultimate sample row 665B and a given penultimate control row 675B.
  • the nitrogen gas passes through bar channels 660 and contacts ink residing in ink row 685, whereby to propel the ink into bar channels 600 (i.e., nitrogen gas passes up bar channel 505 from sample portion 670, whereby to propel ink from ink row 685 up bar channel 505 and into control portion 680) .
  • nitrogen gas is produced by the reaction between hydrogen peroxide and HRP located in penultimate control row 675B, causing nitrogen gas to pass through bar channels 600 and contact ink residing in ink row 685, whereby to propel the ink into bar channels 660 (i.e., nitrogen gas passes down bar channel 505 from control portion 680, whereby to propel ink from ink row 685 down bar channel 505 and into sample portion 670) .
  • the nitrogen gas produced by the control and the nitrogen gas produced by the sample are directed against one another and
  • the ink will move into bar channels 660 (i.e., down bar channels 505) if there is a greater amount of HRP in penultimate control row 675B than there is HRP in penultimate sample row 665B
  • the quantity of the analyte present in the sample can be determined by viewing the direction and distance that the ink travels within bar channels 505.
  • multiplexed volumetric bar chart chip 500 can be used to simultaneously determine (i) whether an analyte is present in a sample at a concentration above a predetermined threshold, and (ii) the quantity of multiple analytes present in a sample, with the quantity of each analyte being indicated in a particular one of a plurality of bar channels 505. See, for example, Figs. 54J-54M which show the test results for various samples.
  • Fig. 54N lists some exemplary analytes and exemplary threshold (i.e., "cutoff") values which may be used when selecting a concentration of HRP to be used for the control.
  • the amount of HRP present in penultimate sample row 665B corresponds to the amount of target analyte present in penultimate sample row 665B
  • the amount of HRP present in penultimate control row 675B corresponds to the amount of HRP introduced into penultimate control row 675B by the user to act as the control for the "competitive" multiplexed volumetric bar chart chip 500.
  • the direction of movement of ink out of ink row 685 and into bar channels 600 is governed by the difference between the amount of nitrogen gas generated by the reaction of the HRP present in penultimate sample row 665B and the amount of nitrogen gas generated by the reaction of the HRP present in penultimate control row 675B. Put another way, the ink will move into bar channels 600 in a direction away from the reaction which produces a greater amount of nitrogen gas (and towards the reaction which produces a lesser amount of nitrogen gas ) .
  • the "competitive" multiplexed volumetric bar chart chip 500 has a wide range and high precision.
  • a difference between a concentration of 1 ⁇ HRP (disposed in one of sample row 665B and control row 675B) and a concentration of 4.5 ⁇ HRP (disposed in the other of sample row 665B and control row 675B) causes ink to move toward the 1 ⁇ HRP side of the multiplexed volumetric bar chart chip.
  • a difference between a concentration of 4 ⁇ HRP (disposed in one of sample row 665B and control row 675B) and a concentration of 5 ⁇ HRP (disposed in the other of sample row 665B and control row 675B) causes ink to move toward the 4 ⁇ HRP side of the multiplexed volumetric bar chart chip.
  • a concentration of 4 ⁇ HRP disposed in one of sample row 665B and control row 675B
  • a concentration of 5 ⁇ HRP disposed in the other of sample row 665B and control row 675B
  • quantity of HRP present in a second sample results in discernably different advancement of the ink into bar channels 600 when offset by the same control.
  • a difference between a concentration of 4 ⁇ HRP in a first sample and 6 ⁇ HRP in a second sample, using a control with a concentration of 5 ⁇ HRP causes a discernable difference in the distance that the ink advances along bar channels 600 (i.e., the ink moves along bar channels 600 toward control row 675B with the second sample and moves toward sample row 665B with the first sample, since the second sample contains a greater amount of HRP than the first sample) .
  • the distance that the ink moves along bar channels 600 is a function of the amount of HRP present in the sample and control, and that the multiplexed volumetric bar chart chip of the present invention accommodates a wide range of sample and control concentrations. It has been found that the ink within ink row 685 moves a discernable distance along bar channels 600 when the concentration of HRP present in sample row 665B (which is a function of the concentration of the analyte present in sample row 665B) is quite high (e.g., > 100 ⁇ HRP), and that the ink within ink row 685 also moves a discernable distance along bar channels 600 when the concentration of HRP present in sample row 665B (which is a function of the concentration of the analyte present in sample row 665B) is quite low
  • control row 675B e.g., in the sub-millimole and nanomole concentration range
  • the difference between a concentration of 500 ⁇ HRP (disposed in one of sample row 665B and control row 675B) and a concentration of 750 ⁇ HRP (disposed in the other of sample row 665B and control row 675B) causes ink to move toward the 500 ⁇ HRP side of multiplexed volumetric bar chart chip 500.
  • the difference between a concentration of 5 nM HRP (disposed in one of sample row 665B and control row 675B) and a concentration of 7.5 nM HRP (disposed in the other of sample row 665B and control row 675B) causes ink to move toward 5 nM HRP side of multiplexed volumetric bar chart chip.
  • novel method and apparatus of the present invention provides instant and visual quantitation of target biomarkers or other molecular analytes and provides a visualized bar chart without the use of instruments, data processing or graphic plotting.
  • the novel method and apparatus of the present invention can be easily used as a point of care determination of the quantity of an analyte (and, preferably, the quantity of multiple analytes) present in a sample. More particularly, the novel method and apparatus of the present invention can be used as a point of care determination of the quantity of
  • a novel multiplexed volumetric bar chart chip is provided which may be used in accordance with the present invention to determine the quantity of a target protein (or other types of biomarkers or other analytes) present in a sample .
  • a novel multiplexed volumetric bar chart chip 700 which is configured to determine the quantity of multiple analytes which may be present in a sample, with the quantity of each analyte which is present in the sample being indicated in a particular one of a plurality of serpentine channels 705.
  • Serpentine channels 705 can be advantageous inasmuch as they provide an increased channel length without increasing the overall size of multiplexed volumetric bar chart chip 700.
  • serpentine channels may be incorporated into
  • multiplexed volumetric bar chart chip 700 In one preferred embodiment of the present invention,
  • multiplexed volumetric bar chart chip 700 comprises three serpentine channels. It should be appreciated that, although serpentine channels 705 are shown as S- shaped channels, serpentine channels 705 may be straight or curved (e.g., circular, z-shaped) or formed in any other configuration which provides a series of channels having a length. As a result of this construction, the review of a particular
  • serpentine channel 705 will indicate the quantity of a particular analyte which may be present in the sample and, significantly, the collective array of the plurality of serpentine channels 705 will
  • volumetric bar chart chip 700 will be discussed in the context of a three-plexed volumetric bar chart chip.
  • multiplexed volumetric bar chart chip 700 comprises two plates, a transparent top plate 710 and a bottom plate 715 (which may or may not be transparent) .
  • top plate 710 (Fig. 54P) has a plurality of sample recesses 720 formed on its bottom surface, with sample recesses 720 being arranged along the upper portion of the bottom surface of top plate 710 so as to provide a plurality of inlets 725 and a plurality of outlets 730 for facilitating loading of a sample into a sample well, as will hereinafter be discussed in greater detail.
  • Top plate 710 also has a plurality of reaction wells 735 formed on its bottom surface, with reaction wells 735 being arranged along the upper portion of the bottom surface of top plate
  • a plurality of inlets 740 and a plurality of outlets 745 are also formed in top plate 710, adjacent reaction wells 735, for permitting loading of a reactant into reaction wells 735 as will hereinafter be discussed in greater detail.
  • a plurality of ink recesses 750 are formed in the bottom surface of top plate 710 and extend
  • An inlet 755 is connected to a far side ink recess 750, and an outlet 760 is formed adjacent to the opposite far side ink recess 750.
  • connection recesses 765 are formed in the bottom surface of top plate 710, disposed intermediate reaction wells 735 and ink recesses 750.
  • each of the serpentine channels 705 are formed on the bottom surface of top plate 710, with each of the serpentine channels 705 extending between (although not in fluid communication with) ink recesses 750 and the bottom edge of top plate 710.
  • Serpentine channels 705 comprise an inlet 770 disposed at the end of each serpentine channel 705 which is adjacent ink recesses 750 and an outlet 775 disposed at the opposite end of each serpentine channel 705 (i.e., adjacent the bottom edge of top plate 710) .
  • sample recesses 720, reaction wells 735, ink recesses 750, serpentine channels 705, inlets 725, 740, 755, 770 and outlets 730, 745, 760, 775 are all formed in the bottom surface of top plate 710 using a conventional etching process of the sort well known in the etching arts.
  • sample recesses 720, reaction wells 735, ink recesses 750, serpentine channels 705, inlets 725, 740, 755, 770 and outlets 730, 745, 760, 775 may be formed in a silicon plate, a plastic plate, a ceramic plate, a quartz plate, a metal oxide plate or other appropriate substrate material.
  • Bottom plate 715 (Fig. 54Q) has a plurality of reactant recesses 780 formed on its top surface, with reactant recesses 780 being arranged such that, when top plate 710 is disposed over bottom plate 715 (i.e., when novel multiplexed volumetric bar chart chip 700 is assembled), reactant recesses 780 fluidically connect inlet 740 and outlet 745 to reaction wells 735
  • Bottom plate 715 also has a plurality of sample wells 785 (Fig. 54Q) formed on its top surface, with sample wells 785 being arranged along the upper portion of the top surface of bottom plate 715.
  • Sample wells 785 are arranged such that, when top plate 710 is disposed over bottom plate 715 (i.e., when novel multiplexed volumetric bar chart chip 700 is assembled), sample recesses 720 connect inlet 725 and outlet 730 to sample wells 785 (Fig. 54R) .
  • Bottom plate 715 also has a plurality of
  • connection recesses 790 (Fig. 54Q) formed on its top surface, with connection recesses 790 being arranged along the upper portion of the top surface of bottom plate 715.
  • Connection recesses 790 are arranged such that, when top plate 710 is disposed over bottom plate 715 (i.e., when novel multiplexed volumetric bar chart chip 700 is assembled), connection recesses 790 fluidically link ink recesses 750 of top plate 710 together (Fig. 54R) , whereby to form a continuous ink row and fluidly connect inlet 755 and outlet 760 as will hereinafter be discussed in greater detail.
  • connection recesses 795 are formed in the top surface of bottom plate 715, disposed below sample wells 785. Connection recesses 795 are arranged such that, when top plate 710 is disposed over bottom plate 715 (i.e., when novel multiplexed volumetric bar chart chip 700 is
  • connection recesses 795 are in fluid communication with ink recesses 750 (Fig. 54S), whereby to permit connection recesses 795 to be filled with liquid ink when ink recesses 750 are filled with liquid ink, as will hereinafter be discussed in greater detail.
  • reactant recesses 780, sample wells 785 and connection recesses 790, 795 are all formed in the top surface of bottom plate 715 using a conventional etching process of the sort well known in the etching arts.
  • reactant recesses 780, sample wells 785 and connection recesses 790, 795 are etched in the top surface of a glass plate.
  • reactant recesses 780, sample wells 785 and connection recesses 790, 795 may be formed in a silicon plate, a plastic plate, a ceramic plate, a quartz plate, a metal oxide plate or other appropriate substrate material.
  • sample recesses 720 in top plate 710 will cooperate with sample wells 785 in bottom plate 715 so as to permit loading of a sample to be assayed into a given sample well 785 via a given inlet 725.
  • Reactant recesses 780 in bottom plate 715 will cooperate with reaction wells 735, inlet 740 and outlet 745 in top plate 710 so as to permit loading of a reactant into a given reaction well 735 via a given inlet 740.
  • Connection recesses 790 will cooperate with ink recesses 750 so as to form a continuous ink row fluidically connecting inlet 755 to outlet 760, whereby to permit loading of a liquid ink (e.g., red ink) into ink recesses 750.
  • a liquid ink e.g., red ink
  • FIG. 54S is a schematic view of top plate 710 and bottom plate 715, with the structures of top plate 710 outlined in solid line and the structures of bottom plate 715 outlined in dashed line.
  • top plate 710 dispositions of recesses 720, 750, 765 and reaction wells 735 in top plate 710, and recesses 780, 790, 795 and sample wells 785 in bottom plate 715, a horizontal slide (to the left) of top plate 710 relative to bottom plate 715 disrupts the aforementioned
  • reaction wells 735 move horizontally relative to sample wells 785, thereby combining reaction wells 735 with sample wells 785, whereby to mix the contents of reaction wells 735 with the contents of sample wells 785, (ii) ink recesses 750 and serpentine channels 705 (and inlet 770 of
  • serpentine channels 705) are shifted horizontally, whereby to align with, and fluidically connect, a given inlet 770 of serpentine channels 705 with a given connection recess 795 of bottom plate 715, and (iii) connection recesses 765 are shifted
  • Fig. 54T is a schematic view of top plate 710 and bottom plate 715, with the structures of top plate 710 outlined in solid line and the
  • bottom plate 715 structures of bottom plate 715 outlined in dashed line, and showing multiplexed volumetric bar chart chip 700 after top plate 710 has been slid
  • connection recess 765 forces liquid ink out of connection recess 795, through inlet 770 and hence, forces liquid ink into serpentine channels 705.
  • the distance that ink travels in a given serpentine channel 705 is proportional to the amount of gas generated by the reaction between the sample and the reactant.
  • multiplexed volumetric bar chart chip 700 can be used to determine the quantity of multiple analytes present in a sample, with the quantity of each specific analyte being indicated in a particular one of the plurality of serpentine channels 705.
  • an analyte-specific antibody is bonded in sample wells 785 of bottom plate 715 of multiplexed volumetric bar chart chip 700.
  • a different analyte-specific antibody is bonded in each sample well 785.
  • Red ink (or some other colored material which is readily discernible through top plate 710 and against bottom plate 715) is introduced into inlet 755 of multiplexed volumetric bar chart chip 700, whereby to fill ink recesses 750
  • connection recesses 795 which are in fluid communication with ink recesses 750 of multiplexed volumetric bar chart chip 700 with red ink.
  • the target analyte i.e., the analyte being tested for
  • the target analyte will bind the analyte-specific antibody which is bound to a sample well 785 of multiplexed volumetric bar chart chip 700, as will hereinafter be discussed in greater detail.
  • the sample is introduced into inlet 725 (Fig. 54R) of multiplexed volumetric bar chart chip 700, so that the sample fills a given sample well 785.
  • sample wells 785 are washed, leaving the target analyte(s) (where present) bound to the analyte-specific antibodies which are, in turn, bound to sample well 785.
  • the amount of the target analyte retained in a given sample well 785 is proportional to the amount of the target analyte present in the sample.
  • a conjugate comprising platinum nanoparticles bound to a detection antibody (which detection
  • the platinum nanoparticle-detection antibody conjugate fills a given sample well 785.
  • the target analyte is present (i.e., where the target analyte is bound to an analyte-specific antibody bound to sample well 785)
  • antibody conjugate will bind the target analyte.
  • sample wells 785 are then washed, leaving only those platinum nanoparticle-detection antibody conjugates which are bound to the target analyte.
  • the amount of platinum nanoparticles present and bound in a given sample well 785 is proportional to the amount of target analyte bound to multiplexed volumetric bar chart chip 700, and hence, proportional to the amount of target analyte present in the sample.
  • hydrogen peroxide from reaction well 735 is thereafter introduced into sample well 785 (via a horizontal slide of top plate 710 relative to bottom plate 715)
  • the reaction between the hydrogen peroxide and the platinum nanoparticles will generate oxygen gas in an amount proportional to the amount of
  • platinum nanoparticles present (and hence,
  • top plate 710 is slid horizontally (Fig. 540) relative to bottom plate 715, causing reaction wells 735 to communicate with sample wells 785, and aligning sample wells 785 with connection recesses 765, which is in turn aligned with, and in fluid communication with, connection recess 795 (which connection recess 795 contains red ink), which is, in turn, aligned with, and in fluid communication with, inlet 760 of serpentine channels 705. See Figs. 54S and 54T.
  • the hydrogen peroxide is introduced to the analyte.
  • the oxygen gas produced passes through connection recess 765, forces liquid ink out of connection recess 795, through inlet 770 and hence, forces liquid ink into serpentine channels 705.
  • the distance that ink travels in a given serpentine channel 705 is proportional to the amount of oxygen gas generated by the reaction between the sample (i.e., the analyte) and the reactant (i.e., the hydrogen peroxide) .
  • a review of a particular serpentine channel 705 indicates the quantity of a given analyte present in a sample .
  • multiplexed volumetric bar chart chip 700 can be used to determine the quantity of multiple analytes present in a sample, with the quantity of each analyte being indicated in a particular one of a plurality of serpentine channels
  • some exemplary analytes may include, but are not limited to, interleukin-1 receptor antagonist (IL-1RA), soluble tumor necrosis factor receptor II (sTNF-RII), and soluble interleukin 1 receptor, type II (IL-1SR2) .
  • IL-1RA interleukin-1 receptor antagonist
  • sTNF-RII soluble tumor necrosis factor receptor II
  • IL-1SR2 soluble interleukin 1 receptor, type II
  • the novel method and apparatus of the present invention can be easily used as a point of care determination of the quantity of an analyte (and, preferably, the quantity of multiple analytes) present in a sample. More particularly, the novel method and apparatus of the present invention can be used as a point of care determination of the quantity of
  • multiplexed volumetric bar chart chip 5 may be
  • one or more biomarkers may be assayed by preparing multiplexed volumetric bar chart chip 5 with biomarker-specific antibodies bound to recesses 30 of row 75B.
  • the biomarkers may comprise one or more from the group consisting of AFP, AFP-L3, DCP, AST, ALT, GGT, CDT, HBcAg, HBeAg, HBsAg, HCV Virus, HbAlC, Ferritin and AFB1.
  • multiplexed volumetric bar chart chip 5 may be
  • one or more biomarkers may be assayed by preparing multiplexed volumetric bar chart chip 5 with biomarker-specific antibodies bound to recesses 30 of row 75B.
  • the biomarkers may comprise one or more from the group consisting of IFN- 2, IFN- ⁇ , IL-l , IL- ⁇ , IL-2, IL-3, IL-6, IL-7, IL-9, IL-12p40, IL-12p70, IL-15, IL-17, TNF- , TNF- ⁇ , IL-4, IL-5, IL-13, IL-10, IL-lra,
  • CXCL1 CXCL1
  • GRO CXCL3
  • IL-8 CXCL8
  • IP-10 CXCL10
  • MCP-1 CCL2
  • MCP-3 CCL7
  • MDC CCL22
  • MIP-la MIP-la
  • CTL3 MIP-lb (CCL4), CSLEX, OPG, OC, PTH, RankL, Adiponectin, EGF, FGF- ⁇ , Flt-3 Ligand, G-CSF, GM-CSF,
  • TGF- TGF- , VEGF and TGF- ⁇ , as well as controls.
  • multiplexed volumetric bar chart chip 5 may be
  • one or more biomarkers may be assayed by preparing multiplexed volumetric bar chart chip 5 with biomarker-specific antibodies bound to recesses 30 of row 75B.
  • the biomarkers may be assayed by preparing multiplexed volumetric bar chart chip 5 with biomarker-specific antibodies bound to recesses 30 of row 75B.
  • the biomarkers may be assayed by preparing multiplexed volumetric bar chart chip 5 with biomarker-specific antibodies bound to recesses 30 of row 75B.
  • the biomarkers may be assayed by preparing multiplexed volumetric bar chart chip 5 with biomarker-specific antibodies bound to recesses 30 of row 75B.
  • the biomarkers may be assayed by preparing multiplexed volumetric bar chart chip 5 with biomarker-specific antibodies bound to recesses 30 of row 75B.
  • the biomarkers may
  • multiplexed volumetric bar chart chip 5 may be
  • biomarkers e.g., biomarkers which are linked to drug abuse
  • Fig. 58 one or more biomarkers (e.g., biomarkers which are linked to drug abuse) may be assayed by preparing multiplexed
  • the biomarkers may be any biomarkers.
  • AMP AMP
  • mAMP mAMP
  • BAR BZO
  • COC MTD
  • OPI OPI
  • PCP PCP
  • THC TCA
  • IgA IgA
  • Creatinine Creatinine
  • multiplexed volumetric bar chart chip 5 may be
  • one or more biomarkers may be assayed by preparing multiplexed volumetric bar chart chip 5 with biomarker-specific antibodies bound to recesses 30 of row 75B.
  • the biomarkers may comprise one or more from the group consisting of ATP, 2,3-DPG and NO.
  • DNA DNA, RNA, And/Or micro-RNA
  • multiplexed volumetric bar chart chip 5 may be
  • DNA, RNA, and/or micro-RNA targets may be assayed by preparing multiplexed volumetric bar chart chip 5 with DNA, RNA, and/or micro-RNA-specific antibodies bound to recesses 30 of row 75B. If desired, a platinum film may be utilized to facilitate the assay. Additional Assays
  • multiplexed volumetric bar chart chip 5 may be
  • biomarkers/materials may comprise one or more from the group consisting of IL-1RA, sTNFRII, IL-1SR2, ATP, 2,3-DPG, hemoglobin, NO and food allergens (e.g., peanut, pine nuts, etc.) .

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Abstract

La présente invention concerne un appareil permettant de déterminer la quantité d'un analyte cible présent dans un échantillon, l'appareil comprenant : une première plaque comprenant une pluralité de cavités agencées de façon à former une pluralité de rangées s'étendant parallèlement l'une à l'autre, et une pluralité de canaux s'étendant perpendiculairement à la pluralité de rangées de la première plaque; et une seconde plaque comprenant une pluralité de cavités agencées de façon à former une pluralité de rangées s'étendant parallèlement l'une à l'autre, et une pluralité de canaux s'étendant perpendiculairement à la pluralité de rangées de la seconde plaque; la première plaque et la seconde plaque étant assemblées ensemble de sorte que la première plaque est positionnée contre la seconde plaque et que les évidements de la première plaque communiquent avec les évidements de la seconde plaque de manière à former une pluralité de rangées d'échantillons, une pluralité de rangées d'échantillons de contrôle, et une rangée d'encre disposée entre la pluralité de rangées d'échantillon et la pluralité de rangées d'échantillons de contrôle.
PCT/US2016/023416 2012-10-16 2016-03-21 Puce graphique à barres volumétriques multiplexées pour biomarqueur hors-laboratoire et/ou quantification d'analyte, comprenant le contrôle compétitif WO2016154113A1 (fr)

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US14/817,258 US10228369B2 (en) 2012-10-16 2015-08-04 Multiplexed volumetric bar chart chip for point of care biomarker and analyte quantitation
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CN108949528A (zh) * 2018-04-10 2018-12-07 南京大学 可视化检测铜、铅、汞离子的多元体积柱芯片及其检测方法
CN108949528B (zh) * 2018-04-10 2022-02-11 南京大学 可视化检测铜、铅、汞离子的多元体积柱芯片及其检测方法
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CN108745426B (zh) * 2018-04-24 2020-08-28 齐齐哈尔医学院 一种用于阿尔茨海默病伴发抑郁症血液相关蛋白检测的微流控芯片及其制备方法和应用
CN113702338A (zh) * 2021-08-27 2021-11-26 深圳大学 一种多通道生物反应传感芯片及其制造方法与装置

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