EP3028045A1 - Device and method for biological analyses - Google Patents
Device and method for biological analysesInfo
- Publication number
- EP3028045A1 EP3028045A1 EP14759031.9A EP14759031A EP3028045A1 EP 3028045 A1 EP3028045 A1 EP 3028045A1 EP 14759031 A EP14759031 A EP 14759031A EP 3028045 A1 EP3028045 A1 EP 3028045A1
- Authority
- EP
- European Patent Office
- Prior art keywords
- analyte
- binding partner
- binding
- detection
- antibody
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Withdrawn
Links
- 238000000034 method Methods 0.000 title claims abstract description 25
- 238000004458 analytical method Methods 0.000 title claims abstract description 12
- 239000012491 analyte Substances 0.000 claims abstract description 186
- 230000027455 binding Effects 0.000 claims abstract description 185
- 238000001514 detection method Methods 0.000 claims description 172
- 238000006243 chemical reaction Methods 0.000 claims description 86
- 239000013641 positive control Substances 0.000 claims description 52
- 239000007788 liquid Substances 0.000 claims description 48
- 108090000765 processed proteins & peptides Proteins 0.000 claims description 47
- 230000015572 biosynthetic process Effects 0.000 claims description 24
- 230000005012 migration Effects 0.000 claims description 20
- 238000013508 migration Methods 0.000 claims description 20
- 239000003550 marker Substances 0.000 claims description 17
- 238000011002 quantification Methods 0.000 claims description 17
- 239000012530 fluid Substances 0.000 claims description 15
- 102000004196 processed proteins & peptides Human genes 0.000 claims description 15
- 238000004891 communication Methods 0.000 claims description 13
- 239000011159 matrix material Substances 0.000 claims description 10
- 101001079904 Homo sapiens Hyaluronan and proteoglycan link protein 1 Proteins 0.000 claims description 4
- 102100028084 Hyaluronan and proteoglycan link protein 1 Human genes 0.000 claims description 4
- 239000011230 binding agent Substances 0.000 claims 1
- 238000012544 monitoring process Methods 0.000 abstract 1
- 239000000523 sample Substances 0.000 description 114
- 238000012360 testing method Methods 0.000 description 48
- 239000000427 antigen Substances 0.000 description 29
- 102000036639 antigens Human genes 0.000 description 29
- 108091007433 antigens Proteins 0.000 description 29
- 238000003018 immunoassay Methods 0.000 description 28
- 239000006249 magnetic particle Substances 0.000 description 27
- 102000013394 Troponin I Human genes 0.000 description 22
- 108010065729 Troponin I Proteins 0.000 description 22
- 102000004169 proteins and genes Human genes 0.000 description 16
- 108090000623 proteins and genes Proteins 0.000 description 16
- 239000002245 particle Substances 0.000 description 15
- 239000000203 mixture Substances 0.000 description 14
- 238000003556 assay Methods 0.000 description 11
- 239000003446 ligand Substances 0.000 description 11
- 230000035945 sensitivity Effects 0.000 description 11
- 239000003153 chemical reaction reagent Substances 0.000 description 10
- 238000005516 engineering process Methods 0.000 description 10
- 102000008394 Immunoglobulin Fragments Human genes 0.000 description 8
- 108010021625 Immunoglobulin Fragments Proteins 0.000 description 8
- 230000003287 optical effect Effects 0.000 description 8
- 239000007787 solid Substances 0.000 description 8
- 241001465754 Metazoa Species 0.000 description 7
- 235000013305 food Nutrition 0.000 description 7
- 230000001900 immune effect Effects 0.000 description 7
- 239000000463 material Substances 0.000 description 7
- 239000000126 substance Substances 0.000 description 7
- 102000004190 Enzymes Human genes 0.000 description 6
- 108090000790 Enzymes Proteins 0.000 description 6
- 102000004903 Troponin Human genes 0.000 description 6
- 108090001027 Troponin Proteins 0.000 description 6
- 229920001184 polypeptide Polymers 0.000 description 6
- 239000002122 magnetic nanoparticle Substances 0.000 description 5
- 239000002105 nanoparticle Substances 0.000 description 5
- 239000013642 negative control Substances 0.000 description 5
- 210000002381 plasma Anatomy 0.000 description 5
- 229920000642 polymer Polymers 0.000 description 5
- 238000001179 sorption measurement Methods 0.000 description 5
- 238000011144 upstream manufacturing Methods 0.000 description 5
- 238000002965 ELISA Methods 0.000 description 4
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 4
- 150000001413 amino acids Chemical class 0.000 description 4
- 239000011324 bead Substances 0.000 description 4
- 230000000747 cardiac effect Effects 0.000 description 4
- 230000008859 change Effects 0.000 description 4
- 238000011534 incubation Methods 0.000 description 4
- 238000002372 labelling Methods 0.000 description 4
- 238000002360 preparation method Methods 0.000 description 4
- 210000002966 serum Anatomy 0.000 description 4
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 3
- 238000010521 absorption reaction Methods 0.000 description 3
- -1 antibodies Proteins 0.000 description 3
- 210000004369 blood Anatomy 0.000 description 3
- 239000008280 blood Substances 0.000 description 3
- 210000004027 cell Anatomy 0.000 description 3
- 150000001875 compounds Chemical class 0.000 description 3
- 238000005859 coupling reaction Methods 0.000 description 3
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 3
- 239000005556 hormone Substances 0.000 description 3
- 229940088597 hormone Drugs 0.000 description 3
- 210000004408 hybridoma Anatomy 0.000 description 3
- 230000001771 impaired effect Effects 0.000 description 3
- 238000012125 lateral flow test Methods 0.000 description 3
- 238000004519 manufacturing process Methods 0.000 description 3
- 230000000849 parathyroid Effects 0.000 description 3
- 239000000243 solution Substances 0.000 description 3
- 238000003786 synthesis reaction Methods 0.000 description 3
- 238000005406 washing Methods 0.000 description 3
- YBJHBAHKTGYVGT-ZKWXMUAHSA-N (+)-Biotin Chemical compound N1C(=O)N[C@@H]2[C@H](CCCCC(=O)O)SC[C@@H]21 YBJHBAHKTGYVGT-ZKWXMUAHSA-N 0.000 description 2
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 2
- 101000945318 Homo sapiens Calponin-1 Proteins 0.000 description 2
- 101000652736 Homo sapiens Transgelin Proteins 0.000 description 2
- 102000003982 Parathyroid hormone Human genes 0.000 description 2
- 108090000445 Parathyroid hormone Proteins 0.000 description 2
- 102100031013 Transgelin Human genes 0.000 description 2
- 102100036859 Troponin I, cardiac muscle Human genes 0.000 description 2
- 101710128251 Troponin I, cardiac muscle Proteins 0.000 description 2
- 230000003321 amplification Effects 0.000 description 2
- 239000012472 biological sample Substances 0.000 description 2
- 229940098773 bovine serum albumin Drugs 0.000 description 2
- 230000008878 coupling Effects 0.000 description 2
- 238000010168 coupling process Methods 0.000 description 2
- 230000002950 deficient Effects 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 230000002255 enzymatic effect Effects 0.000 description 2
- 238000004128 high performance liquid chromatography Methods 0.000 description 2
- 230000002163 immunogen Effects 0.000 description 2
- 239000004816 latex Substances 0.000 description 2
- 229920000126 latex Polymers 0.000 description 2
- 239000012528 membrane Substances 0.000 description 2
- 238000003199 nucleic acid amplification method Methods 0.000 description 2
- 239000000199 parathyroid hormone Substances 0.000 description 2
- 229960001319 parathyroid hormone Drugs 0.000 description 2
- 238000000746 purification Methods 0.000 description 2
- 230000002285 radioactive effect Effects 0.000 description 2
- 239000011780 sodium chloride Substances 0.000 description 2
- 239000007790 solid phase Substances 0.000 description 2
- 239000000758 substrate Substances 0.000 description 2
- JJAHTWIKCUJRDK-UHFFFAOYSA-N succinimidyl 4-(N-maleimidomethyl)cyclohexane-1-carboxylate Chemical compound C1CC(CN2C(C=CC2=O)=O)CCC1C(=O)ON1C(=O)CCC1=O JJAHTWIKCUJRDK-UHFFFAOYSA-N 0.000 description 2
- 238000012546 transfer Methods 0.000 description 2
- 210000002700 urine Anatomy 0.000 description 2
- 238000012800 visualization Methods 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 102100031126 6-phosphogluconolactonase Human genes 0.000 description 1
- 108010029731 6-phosphogluconolactonase Proteins 0.000 description 1
- 241000251468 Actinopterygii Species 0.000 description 1
- 102000002260 Alkaline Phosphatase Human genes 0.000 description 1
- 108020004774 Alkaline Phosphatase Proteins 0.000 description 1
- 208000034656 Contusions Diseases 0.000 description 1
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 1
- 238000012286 ELISA Assay Methods 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- 101001121580 Enterobacteria phage PRD1 Adsorption protein P2 Proteins 0.000 description 1
- 208000035126 Facies Diseases 0.000 description 1
- PDLGMYVCPJOYAR-DKIMLUQUSA-N Glu-Leu-Phe-Ala Chemical compound OC(=O)CC[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@H](C(=O)N[C@@H](C)C(O)=O)CC1=CC=CC=C1 PDLGMYVCPJOYAR-DKIMLUQUSA-N 0.000 description 1
- 108010018962 Glucosephosphate Dehydrogenase Proteins 0.000 description 1
- 108010001336 Horseradish Peroxidase Proteins 0.000 description 1
- 102000017727 Immunoglobulin Variable Region Human genes 0.000 description 1
- 108010067060 Immunoglobulin Variable Region Proteins 0.000 description 1
- 102000004856 Lectins Human genes 0.000 description 1
- 108090001090 Lectins Proteins 0.000 description 1
- 241001529936 Murinae Species 0.000 description 1
- 239000000020 Nitrocellulose Substances 0.000 description 1
- 101001125164 Parietaria judaica Probable non-specific lipid-transfer protein 2 Proteins 0.000 description 1
- 206010035226 Plasma cell myeloma Diseases 0.000 description 1
- 239000004793 Polystyrene Substances 0.000 description 1
- 101000580771 Pseudomonas phage phi6 RNA-directed RNA polymerase Proteins 0.000 description 1
- 101001121571 Rice tungro bacilliform virus (isolate Philippines) Protein P2 Proteins 0.000 description 1
- 108010090804 Streptavidin Proteins 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 239000002313 adhesive film Substances 0.000 description 1
- 238000001042 affinity chromatography Methods 0.000 description 1
- 239000000956 alloy Substances 0.000 description 1
- 229910045601 alloy Inorganic materials 0.000 description 1
- 150000001412 amines Chemical class 0.000 description 1
- 210000003567 ascitic fluid Anatomy 0.000 description 1
- 150000001540 azides Chemical class 0.000 description 1
- 210000003719 b-lymphocyte Anatomy 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 102000005936 beta-Galactosidase Human genes 0.000 description 1
- 108010005774 beta-Galactosidase Proteins 0.000 description 1
- 238000011953 bioanalysis Methods 0.000 description 1
- 238000004166 bioassay Methods 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 239000013060 biological fluid Substances 0.000 description 1
- 229960002685 biotin Drugs 0.000 description 1
- 235000020958 biotin Nutrition 0.000 description 1
- 239000011616 biotin Substances 0.000 description 1
- 238000011088 calibration curve Methods 0.000 description 1
- 210000001175 cerebrospinal fluid Anatomy 0.000 description 1
- 235000013351 cheese Nutrition 0.000 description 1
- 238000007385 chemical modification Methods 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- 238000004737 colorimetric analysis Methods 0.000 description 1
- 239000000470 constituent Substances 0.000 description 1
- UFULAYFCSOUIOV-UHFFFAOYSA-N cysteamine Chemical compound NCCS UFULAYFCSOUIOV-UHFFFAOYSA-N 0.000 description 1
- 239000002274 desiccant Substances 0.000 description 1
- 230000001627 detrimental effect Effects 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- 238000000502 dialysis Methods 0.000 description 1
- 230000004069 differentiation Effects 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 238000010494 dissociation reaction Methods 0.000 description 1
- 230000005593 dissociations Effects 0.000 description 1
- 231100000673 dose–response relationship Toxicity 0.000 description 1
- 239000000975 dye Substances 0.000 description 1
- 235000013601 eggs Nutrition 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- MGHUAQWFVMJEFI-UHFFFAOYSA-N ethane-1,1-dithiol 2,2,2-trifluoroacetic acid tri(propan-2-yl)silane hydrate Chemical compound O.C(C)(C)[SiH](C(C)C)C(C)C.C(C)(S)S.FC(C(=O)O)(F)F MGHUAQWFVMJEFI-UHFFFAOYSA-N 0.000 description 1
- 230000001747 exhibiting effect Effects 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 235000019688 fish Nutrition 0.000 description 1
- 239000012634 fragment Substances 0.000 description 1
- 238000004108 freeze drying Methods 0.000 description 1
- 235000015203 fruit juice Nutrition 0.000 description 1
- 238000010353 genetic engineering Methods 0.000 description 1
- 239000003365 glass fiber Substances 0.000 description 1
- 239000010931 gold Substances 0.000 description 1
- 229910052737 gold Inorganic materials 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 239000008241 heterogeneous mixture Substances 0.000 description 1
- 230000003053 immunization Effects 0.000 description 1
- 238000002649 immunization Methods 0.000 description 1
- 238000003119 immunoblot Methods 0.000 description 1
- 238000003317 immunochromatography Methods 0.000 description 1
- 238000010166 immunofluorescence Methods 0.000 description 1
- 239000002523 lectin Substances 0.000 description 1
- 238000000670 ligand binding assay Methods 0.000 description 1
- 238000004020 luminiscence type Methods 0.000 description 1
- 210000004698 lymphocyte Anatomy 0.000 description 1
- 238000004949 mass spectrometry Methods 0.000 description 1
- 239000008268 mayonnaise Substances 0.000 description 1
- 235000010746 mayonnaise Nutrition 0.000 description 1
- 235000012054 meals Nutrition 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 235000013372 meat Nutrition 0.000 description 1
- 238000005374 membrane filtration Methods 0.000 description 1
- 239000002923 metal particle Substances 0.000 description 1
- 235000013336 milk Nutrition 0.000 description 1
- 239000008267 milk Substances 0.000 description 1
- 210000004080 milk Anatomy 0.000 description 1
- 238000011512 multiplexed immunoassay Methods 0.000 description 1
- 201000000050 myeloid neoplasm Diseases 0.000 description 1
- 208000010125 myocardial infarction Diseases 0.000 description 1
- 229920001220 nitrocellulos Polymers 0.000 description 1
- 230000007170 pathology Effects 0.000 description 1
- 238000010647 peptide synthesis reaction Methods 0.000 description 1
- 239000004033 plastic Substances 0.000 description 1
- 210000004910 pleural fluid Anatomy 0.000 description 1
- 238000012123 point-of-care testing Methods 0.000 description 1
- 229920000728 polyester Polymers 0.000 description 1
- 229920002223 polystyrene Polymers 0.000 description 1
- 238000001556 precipitation Methods 0.000 description 1
- 239000002243 precursor Substances 0.000 description 1
- 238000009597 pregnancy test Methods 0.000 description 1
- 238000007639 printing Methods 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 238000009877 rendering Methods 0.000 description 1
- 210000003296 saliva Anatomy 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 238000003118 sandwich ELISA Methods 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 239000010802 sludge Substances 0.000 description 1
- VUFNRPJNRFOTGK-UHFFFAOYSA-M sodium;1-[4-[(2,5-dioxopyrrol-1-yl)methyl]cyclohexanecarbonyl]oxy-2,5-dioxopyrrolidine-3-sulfonate Chemical compound [Na+].O=C1C(S(=O)(=O)[O-])CC(=O)N1OC(=O)C1CCC(CN2C(C=CC2=O)=O)CC1 VUFNRPJNRFOTGK-UHFFFAOYSA-M 0.000 description 1
- 239000002689 soil Substances 0.000 description 1
- 238000000638 solvent extraction Methods 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 230000009870 specific binding Effects 0.000 description 1
- 230000001629 suppression Effects 0.000 description 1
- 235000013311 vegetables Nutrition 0.000 description 1
- 230000000007 visual effect Effects 0.000 description 1
- 238000001262 western blot Methods 0.000 description 1
- 235000013618 yogurt Nutrition 0.000 description 1
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
- G01N33/54386—Analytical elements
-
- 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
- G01N33/54386—Analytical elements
- G01N33/54387—Immunochromatographic test strips
- G01N33/54388—Immunochromatographic test strips based on lateral flow
- G01N33/54389—Immunochromatographic test strips based on lateral flow with bidirectional or multidirectional lateral flow, e.g. wherein the sample flows from a single, common sample application point into multiple strips, lanes or zones
-
- 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
- G01N33/558—Immunoassay; Biospecific binding assay; Materials therefor using diffusion or migration of antigen or antibody
Definitions
- the present invention relates to the field of the detection and / or quantification of analyte (s) in liquid samples during a biological analysis.
- the present invention particularly relates to a device and a method for performing a sandwich-type immunoassay for detecting and / or quantifying at least one analyte in a sample.
- the invention allows the biological control (control of the functional / operational character) of the analyte binding partners used in the context of said sandwich-type immunoassay, in order to avoid obtaining false-negative results when all or part of the connecting partners used are defective.
- Sandwich type immunoassays are widely used in bioanalysis to detect and / or quantify a given analyte. These sandwich type immunoassays (or more simply “sandwich immunoassays”) use a first binding partner, called “capture binding partner”, generally immobilized on a solid phase to specifically bind the desired analyte, and a second binding partner, called “detection binding partner”, labeled and also intended to bind specifically with the desired analyte, thereby revealing the binding between the capture binding partner and the analyte, and thus the presence of the analyte.
- capture binding partner generally immobilized on a solid phase
- detection binding partner labeled and also intended to bind specifically with the desired analyte, thereby revealing the binding between the capture binding partner and the analyte, and thus the presence of the analyte.
- the desired analyte is "sandwiched" between said first and second binding partners, the first binding partner ("capture") generally being present in excess of the desired analyte.
- the capture binding partner may, for example, be immobilized on a solid support (by covalent bonding, adsorption or any other suitable method) and the amount must generally be such that the number of binding sites present on the binding partner of the capture is greater than the number of antigen molecules present in the standard or unknown solutions.
- the labeled detection binding partner may, for example, be added either simultaneously or after a first incubation and washing, and binds to the antigen, previously immobilized to the capture binding partner.
- a simple wash separates the binding partner-capture-analyte-detection binding partner (labeled) partners from the free labeled detection binding partners present in excess.
- labeled detection binding partner In order for the labeled detection binding partner to bind to the analyte already engaged in a reaction with the capture binding partner, it is necessary for both binding partners to recognize different areas of the analyte.
- the labeled detection binding partner may be, for example, via a radioactive isotope or enzymatically.
- the signal corresponding to the capture binding partner-analyte-labeled detection binding partner-partner complex is a signal representative of the amount of analyte (s) present (s) in the sample tested.
- the capture and detection binding partners are antibodies recognizing epitopes different from the desired analyte, which in this embodiment consists of an antigen.
- the antibodies used as capture (“first binding partner”) and detection (“second binding partner”) binding partners may be, for example, multivalent polyclonal antibodies or different specific monoclonal antibodies.
- Sandwich type immunoassays because of their high sensitivity, can serve as a basis for different types of tests.
- EIA Enzymatic immunoassays
- the ELISA test (of the English “Enzyme-linked immunosorbent assay", literally “bound enzyme immunoabsorption assay”) sandwich.
- this test is as follows: a. a surface (for example a plate, such as a "96-well” plate) is
- the sample likely to contain the desired antigen is added, so that any antigen present binds to the capture antibody;
- the so-called “detection” antibody conjugated directly or indirectly with an enzyme, is added and binds to the antigen, thus forming a capture antibody-antigen-detection antibody complex;
- the plate is rinsed so as to separate this complex from the free labeled detection antibodies present in excess;
- the chromogenic or fluorogenic enzymatic substrate (referred to as the "ELFA” assay) is added; the latter is converted by the enzyme into a detectable form (colored or fluorescent).
- the sandwich-type immunoassays can advantageously be implemented using the "Magnotech” technology from Philips.
- a test of this type namely using the “Magnotech” technology, is described in particular in the publication by Bruis et al. [1].
- This technology uses a reaction cartridge, preferably made of plastic material, comprising a zone for applying a liquid sample and one or more reaction chambers comprising the various reagents necessary for the test, this reaction zone being connected to the zone of applying the sample by a channel allowing the capillary flow of the liquid sample from the application zone to said reaction zone.
- the reaction zone comprises in particular magnetic nanoparticles (of the order of a hundred nanometers in diameter, for example superparamagnetic particles) dried and coated with an antibody of interest capable of binding specifically to a first epitope of the analyte sought.
- said reaction zone comprises, at a region that may be qualified as a "detection region", antibodies capable of specifically recognizing a second epitope of the desired antigen.
- this reaction cartridge may be disposed within a portable device comprising two electromagnets positioned above and below the reaction cartridge. The result of the immunoassay is read after magnetic actuation, the upper electromagnet serving as the washing magnet, while the lower electromagnet acts as a connecting magnet.
- This technique also makes it possible to quantify the desired analyte by establishing a calibration curve from the variation of the optical signal as a function of the concentration of magnetic nanoparticles and thus indirectly of the analyte.
- sandwich type immunoassays are implemented in unit tests, for example in rapid unit tests, such as rapid screening tests (RDTs).
- RDTs rapid screening tests
- the latter are generally in the form of a "cassette", like pregnancy tests, and are relatively easy to use.
- These tests are generally based on the principle of immunochromatography or membrane filtration: the sample (for example whole blood, serum, urine) deposited on the support will, for example, migrate by capillary action. bringing with it reagents already present then will meet binding partners deposited on the membrane when the sample is filtered by this membrane.
- These rapid unit tests are also referred to as “lateral flow assays” or “lateral flow tests” and have been extensively described in the literature (see for example Yager et al [5] or Wild [6]).
- side-flow testing devices can be used to determine the presence of many types of analytes, such as antibodies, antigens, hormones, proteins, chemical molecules, present in liquid samples.
- analytes such as antibodies, antigens, hormones, proteins, chemical molecules
- the analyte to be detected if it is present in the liquid sample deposited at the area of application, binds to a labeled detection binding partner.
- the complexes thus formed then migrate to the reaction zone where they are immobilized at the capture zone by binding with a capture binding partner.
- the user can note the presence of the desired analyte by demonstrating a detectable signal, determined according to the type of marker associated with the detection binding partner.
- the presence of the analyte of interest in the sample is materialized as a detectable line, usually referred to as a test line.
- the reaction zone generally comprises a sample migration control region that will indicate to the user that at least a portion of the sample has passed through the matrix, upstream of the control region, and particularly at the catchment area. This can be, for example, by the revelation of a control line of a predetermined color.
- the patent application WO 2012/066235 discloses a device for performing a rapid unit test, which device incorporates a positive control.
- a control region is provided within this device, for example downstream of the viewing area of the results, or in parallel with the latter, which control region comprises at least one analogue of the desired analyte capable of bind to the labeled detection binding partner.
- the fast unitary test object of the application WO 2012/066235 allows not only the detection of the desired analyte but also to test the functional / operational character of the detection link partner.
- this control is negative, the result of this test is considered uninterpretable and the test must be renewed.
- the Applicant discovered, against all odds, that the functionality (functional / operational character) of the capture binding partner, generally immobilized / adsorbed on a solid surface, could be affected in whole or in part by external factors, such as light, temperature, etc.
- this generates "false negatives", in the the analyte, although present in the tested sample, is not detected in the so-called “visualization of results” zone, but the control region makes it possible to confirm the functionality (functional / operational character) of the detection antibody, and therefore the apparent reliability of the test, while the capture partner is failing. The user / operator therefore incorrectly concludes that the desired analyte is not present in the sample tested.
- an object of the present invention relates to a device for detecting and / or quantifying at least one analyte in a liquid sample during a biological analysis using at least two PI and P2 binding partners of said analyte, the first PI binding partner being immobilized within said device and the second P2 binding partner being immobilizable by forming a sandwich-type complex with the analyte and the first PI binding partner, said device comprising at least two zones fluid communication, namely: a) an application zone of the liquid sample, and
- reaction zone b) comprising at least a first analogue CTRL1 of said analyte, said first analogue CTRL1 being immobilizable by binding to the first immobilized PI binding partner, and at least one second analog CTRL2 of the analyte, said second analog CTRL2 being immobilized within said device so as to bind to the second immobilizable link partner P2 and thus immobilize the latter
- said reaction zone b) comprising at least the following three regions: bl ) a first region for detecting and / or quantifying the analyte, by demonstrating the formation of a sandwich-type complex between the first PI binding partner, the analyte and the second P2 binding partner, b.2) a second biological control region (functional / operational control) of the first link partner PI, at which the first link partner PI is immobilized, the detection of a link between the first partner of
- the device according to the invention comprises a migration zone of the liquid sample c), allowing the migration of the liquid sample from the zone of application of the liquid sample a) to the zone reactional b).
- the presence of this migration zone of the liquid sample c) is particularly desirable when the device according to the invention is suitable for use in a fast unitary test.
- the migration zone of the liquid sample c) comprises a matrix.
- the first and second PI and P2 binding partners consist of antibodies and the first and second analogs of the analyte CRTL1 and CRTL2 comprise at least the epitopes recognized respectively by the first and second PI and P2 antibodies.
- the first and second analogs of the analyte CRTL1 and CRTL2 are peptides containing at least the epitopes recognized respectively by the first and second antibodies PI and P2.
- the second binding partner P2 and the first analog of the analyte CTRL1 are initially deposited - advantageously deposited and dried - in the device and are adapted to be resuspended in the presence of the liquid sample.
- the reaction zone b) of the device according to the invention comprises at least two distinct parts, for example two distinct reaction chambers, without direct communication of fluid between the two parts, the first part comprising the first region bl) and the second part comprising the second and third regions b.2) and b.3).
- the first part comprises, in addition to the first immobilized PI binding partner, the second immobilizable P2 binding partner and the second part comprises, in addition to the first PI binding partner and the second analog of the immobilized analyte CRTL2, the first analogue of the analyte CTRL1 and the second link partner P2 immobilizable.
- the reaction zone b) of the device according to the invention comprises at least two distinct parts, for example two distinct reaction chambers, without direct fluid communication between the two parts, the first part comprising the first and third regions bl) and b.3) and the second portion comprising the second region b.2).
- the first part comprises, besides the first PI binding partner and the second analog of the immobilized analyte CRTL2, the second immobilizable P2 binding partner and the second part comprises, besides the first immobilized PI binding partner, the first analog of the immobilized analyte CRTL2.
- the reaction zone b) of the device according to the invention comprises at least three distinct parts, for example three reaction chambers, without fluid communication between the three distinct parts, the first part comprising the region b1), the second part comprising region b.2) and the third part comprising region b.3).
- the first part comprises, in addition to the first immobilized PI binding partner, the second immobilizable P2 binding partner
- the second part comprises, in addition to the second analogue of the immobilized analyte CTRL2
- the second third part comprises, besides the first immobilized PI binding partner, the first analogue of the immobilizable analyte CTRL1.
- the reaction zone b) of the device according to the invention comprises at least one part, for example a reaction chamber, said part comprising:
- the first analogue of the analyte CTRL1 is associated, directly or indirectly, with an M1 marker making it possible to demonstrate the binding of said first analogue of the analyte CTRL1 with the first binding partner P1 at the level of the regions b1) and b). 2), and / or
- the second P2 binding partner being associated, directly or indirectly, with a marker M2, different from the marker M1, making it possible to demonstrate the formation of the sandwich-type complex between the second binding partner P2, the analyte and the first partner PI link at the level of regions b1 and b.2),
- the zone of application of the liquid sample a comprises a filter.
- Another subject of the invention relates to a method for detecting and / or quantifying at least one analyte in a liquid sample during a biological analysis using at least two binding partners of the PI and P2 analyte, said method comprising the steps of:
- the invention also relates to the use of at least two analogs of an analyte CTRL1 and CTRL2 for the biological control (functional / operational control), respectively, of at least two PI and P2 binding partners, said partners PI and P2 binding system for detecting and / or quantifying said analyte by demonstrating the formation of a sandwich-type complex between the second P2 binding partner, the analyte and the first binding partner P1.
- detecting ... at least one analyte it is intended to detect the presence of the desired analyte within the liquid sample of interest.
- sample By “quantifying at least one analyte”, it is meant to determine the amount (e.g. concentration) / assaying the desired analyte within said liquid sample.
- sample By “sample” is meant a small or small amount isolated from an entity for analysis. It can be a sample of clinical origin, human or animal, from a biological fluid sample, or a food sample, from any type of food, or a sample of the production or processing environment of food. As previously indicated, this sample is liquid.
- sample of clinical origin is meant a sample taken from a patient or individual (human or animal), and may contain an analyte as defined below.
- This sample may be in particular a liquid biological sample such as a sample of blood, serum, plasma, saliva, urine, cerebrospinal fluid, pleural fluid, peritoneal fluid (non-exhaustive list).
- sample of food origin there may be mentioned, for example, a food sample of water, drink such as milk or fruit juice, yoghurt, meat, eggs, vegetables , mayonnaise, cheese, fish, etc.
- This food-based sample may also be derived from a feed intended for animals, such as in particular a sample derived from animal meal.
- sample of environmental origin there may be mentioned, for illustrative purposes, a sample for surface control or water or taken from effluents, sludge, soil, plants, etc. The sample taken can be used as it is, or prior to the biological analysis, undergo an enrichment-type preparation, dilution, extraction, concentration, purification, according to methods known to those skilled in the art.
- liquid sample By “liquid” sample, it is appropriate to understand the samples taken directly in liquid form but also semi-solid or solid samples to the extent that they can be converted into a liquid sample by any appropriate method known to those skilled in the art. Of course, when the sample is solid or semi-solid, it must be pre-processed to be transformed into a liquid sample. This sample is available by any type of sampling known to those skilled in the art.
- analyte should be understood in a broad sense to mean a chemical, biological or biochemical substance that is the subject of one or more analysis (s).
- analytes there may be mentioned an antigen, an antibody, a hormone, a protein or a chemical molecule (non-exhaustive list).
- the nature of the analyte sought will dictate the nature of the binding partners to be used.
- binding partners such as receptors, antibodies, antibody fragments, antibody analogs and any other ligand capable of bind to a protein or an antigen.
- antibody analogs are meant biological and / or chemical compounds which possess the same binding capacities as antibodies or antibody fragments or similar binding capacities.
- the antibody analogues include small proteins which, like the antibodies, are able to bind to a biological target, thus allowing it to be detected, captured or simply targeted within an organism or organism. a biological sample.
- analogue of the analyte is meant a chemical, biological or biochemical species which has physicochemical and biological properties (for example in terms of affinity for a given ligand) close to those of the analyte.
- the analogue of the analyte is a protein, such as an antibody, a mixture of proteins (for example a mixture of antibodies), a polypeptide, a mixture of polypeptides, a peptide, a mixture of peptides, an antibody fragment, a mixture of antibody fragments, an antibody analog, a mixture of antibody analogs, or combinations thereof.
- the binding partner is an antibody, an antibody fragment, or an antibody analog
- the analyte is a protein, a polypeptide, or a peptide
- the analyte analog is also a protein, a polypeptide or peptide.
- parts ... without direct fluid communication must be understood, within the meaning of the present application, as designating parts (for example reaction chambers) adapted to receive a fluid, in this case a liquid, said the parts being further adapted so that, once this liquid received, the latter can not freely communicate between said parts (for example between said reaction chambers).
- the absence of direct fluid communication can be ensured, for example, by the partitioning of the parts (for example reaction chambers) by means of sealed septum (s) and, more generally, any known technical solution of the invention. skilled person.
- the device according to the invention has a very good sensitivity, namely that the probability of obtaining a "false negative” result is extremely low. almost zero or no.
- sensitivity is meant the ability to give a positive result when the desired analyte is present in the tested liquid sample.
- the biological analysis implemented using the device according to the invention covers any immunoassay using at least two binding partners such as a sandwich type immunoassay (also called a sandwich type immunoassay).
- a sandwich type immunoassay also called a sandwich type immunoassay.
- immunoassay also called a sandwich type immunoassay.
- the prefix "immuno" in the term “immunoassay” is not to be considered in the present application as strictly indicating that the binding partner is necessarily a partner of immunological origin, such as an antibody or an antibody fragment.
- this term is more widely used to designate tests and processes in which the binding partner is not a partner of origin / immunological nature but consists, by for example, an analyte receptor that one wishes to detect and / or quantify.
- the binding partner concerned is able to bind to the desired analyte, preferably in a specific manner.
- ligand binding assay which could be translated into French as "assay using binding. to a ligand ", while the term” immuno "is included in the verbatim title corresponding to the acronym ELISA.
- immuno is used in this application to refer to any biological assay using at least one binding partner adapted to bind to the analyte of interest and detect and / or quantify the latter. preferably, specifically, even when said binding partner is not of a strictly immunological nature or origin.
- the first PI binding partner and the second P2 binding partner are selected, for example, from the group consisting of antibody, antibody mixture, antibody fragment, antibody fragment mixture, antibody analogue, mixture of Antibody analogs, antigen, antigen mixture, protein, protein mixture, polypeptide, polypeptide mixture, peptide, peptide mixture.
- the first PI binding partner is a "capture binding partner” insofar as it is immobilized, directly or indirectly, on a solid surface.
- the second link partner P2 is preferably a "detection link partner” and is associated - directly or indirectly - with a detection element making it possible to demonstrate the presence of this second P2 link partner and, most importantly, any and all elements (analyte (s), reagent (s), etc.) related to it.
- This detection element may be a marker reagent but, more broadly, any element making it possible to demonstrate the presence of this second P2 binding partner and, especially, any / all element (s) (analyte (s), reagent (s), etc.) related to it.
- the nanoparticle associated with the detection binding partner acts as a detection element, insofar as the attenuation of the reflected light beam due to this nanoparticle is indicative of the presence of the detection antibody at the level of the detection region and, consequently, the formation of a sandwich-type complex between the detection antibody, the desired analyte and the capture antibody.
- the detection element is a marker reagent capable of directly or indirectly generating a detectable signal.
- these marker reagents may consist of:
- enzymes which produce a detectable signal for example by colorimetry, fluorescence, luminescence, such as horseradish peroxidase, alkaline phosphatase, ⁇ -galactosidase, glucose-6-phosphate dehydrogenase,
- chromophores such as fluorescent compounds, luminescent compounds, dyes,
- fluorescent molecules such as Alexa or phycocyanines, radioactive molecules such as 32 P, 35 S or 125 I, metal particles or alloys, such as colloidal gold particles,
- polymer particles such as colored latex particles.
- Indirect detection systems can also be used, for example ligands capable of reacting with an anti-ligand.
- the ligand / anti-ligand pairs are well known to those skilled in the art, which is the case, for example, of the following pairs: biotin / streptavidin, hapten / antibody, antigen / antibody, peptide / antibody, sugar / lectin, molecule / receiver. In this case, it is the ligand that carries the binding partner.
- the anti-ligand can be detectable directly by the aforementioned marker reagents or be itself detectable by a ligand / anti-ligand.
- Demonstration of the formation of the sandwich-type complex between the first PI binding partner and the second P2 binding partner may be carried out by any technique known to those skilled in the art, in particular by direct visualization of a reaction due to marker reagent associated with the second P2 binding partner or else by means of detection means, for example of the optical type, as is the case under the "Magnotech" operating protocol.
- At least one of the first and second binding partners PI and P2 - preferably both - are of nature and / or of immunological origin.
- at least one of said PI and P2 binding partners - preferably both - are antibodies.
- antibodies is meant a polyclonal antibody, a monoclonal antibody, a humanized antibody, a human antibody or a fragment of said antibodies, in particular the Fab, Fab ', F (ab') 2, ScFv, Fv fragments, fd.
- said antibodies should be specific for the desired analyte, i.e., as far as possible, they do not cross-react with other analytes; the antibodies with the highest affinities and the weakest dissociation constants being the preferred antibodies for the purposes of the present invention.
- the polyclonal antibodies can be obtained by immunization of an animal with the appropriate immunogen, followed by the recovery of the desired antibodies in purified form, by removal of the serum of said animal, and separation of said antibodies from the other constituents of the serum, in particular by chromatography. affinity on a column on which is immobilized an analyte (antigen) specifically recognized by the antibodies.
- the monoclonal antibodies can be obtained by the hybridoma technique whose general principle is recalled below.
- an animal is immunized with the appropriate immunogen whose B cells are then able to produce antibodies against this antigen.
- These antibody-producing lymphocytes are then fused with "immortal" (usually murine) myeloma cells for give rise to hybridomas.
- the cells capable of producing a particular antibody and of multiplying indefinitely are then selected.
- Each hybridoma is multiplied in clone form, each leading to the production of a monoclonal antibody whose recognition properties vis-à-vis the desired antigen can be tested for example by ELISA, by immunoblotting (Western blot) in one or two dimensions, in immunofluorescence, or with a biosensor.
- the monoclonal antibodies thus selected are subsequently purified, for example by affinity chromatography.
- Monoclonal antibodies can also be recombinant antibodies obtained by genetic engineering, by techniques well known to those skilled in the art.
- the desired analyte is referred to as "antigen”
- the first and second PI and P2 binding partners recognize two distinct epitopes - respectively called E1 and E2 - of this antigen.
- PI is preferably a "capture antibody”, as long as it is immobilized, directly or indirectly, on a solid surface.
- the antibody P2 is preferably a "detection antibody” and is associated - directly or indirectly - with a detection element making it possible to demonstrate the presence of this second antibody P2 and, above all, of all / all elements ( s) (analyte (s), reagent (s), etc.) related thereto.
- This detection element is as defined above.
- immobilized binding partner means that the binding partner is attached directly or indirectly within the device by any method known to those skilled in the art such as by adsorption or covalent bonding.
- peptide containing an epitope recognized by an antibody refers to any sequence of amino acids comprising at least the epitope recognized by the binding partner concerned.
- the peptide may be limited to the epitope itself, or include a few additional amino acids, until the protein is obtained, as long as the peptide retains recognition activity by the relevant binding partner.
- the peptide may also contain one or more mutated amino acids, since, again, the peptide retains the recognition activity by the relevant binding partner.
- a capture partner pair also referred to as a capture binding partner
- a detection partner also referred to as a detection binding partner
- the peptide may comprise the E1 epitope of the first capture partner and the E2 epitope of the other capture partner.
- the term "matrix" refers to any type of material capable of ensuring the flow and transfer of a liquid. The transfer of the liquid can be ensured by capillary force. This is particularly advantageous when the device according to the invention is a device for implementing a lateral flow assay.
- the matrix may be, for example, at least one material bibulous. Bibulous materials are materials that easily absorb a liquid and through which the liquid is transported by capillarity. As a crumbly material, there may be mentioned, for example, nitrocellulose, polyester, glass fibers, etc.
- FIG. 1 is a schematic representation of the concept underlying the present invention
- FIG. 2 is a view from above of a reaction cartridge that can be used in an "Magnotech” type opto-magnetic immunoassay device;
- FIGS. 3A, 3B, 3C and 3D illustrate the operation of an opto-magnetic immunoassay technology platform such as the "Magnotech” platform;
- FIG. 4 is a view from above of a device according to a first embodiment of a first aspect of the invention.
- FIG. 5 schematically illustrates a first configuration of this first embodiment of the invention
- FIG. 6 schematically represents a second configuration of this first embodiment of the invention
- FIG. 7 is a view from above of a device according to a second embodiment of a first aspect of the invention.
- FIG. 9 schematically represents a third embodiment of a first aspect of the invention.
- FIGS. 10A, 10B, 10C and 10D represent top views of a lateral flow test as described in application WO 2012/066235, in the name of the Applicant;
- FIGS. 11A, 11B, 11C, 11D, 11E and 11F are diagrammatic views from above of a device according to a second aspect of the invention, suitable for implementing a unilateral flow measurement. ;
- FIG. 12 is a schematic representation of the biological control of the P2 binding partner in the context of the detection of troponin I (TnI) as an analyte, PI and P2 being anti-TnI monoclonal antibodies, respectively 19C7 and 560. , and CTRL2 containing an epitope peptide of P2;
- TnI troponin I
- FIG. 13 is a schematic representation of a Magnotech cartridge having two reaction chambers, in which the P2 binding partner is biologically controlled, the chamber 1 being coated with two spots, each for the detection of Troponin I (spot PI on which is immobilized the anti-Troponin I 19C7 monoclonal antibody (PI binding partner)), and the chamber 2 is coated with three spots, one of the spots being for the detection of Troponin I (spot PI, identical to the spots PI of chamber 1) and the other two (spot CTRL2-1 and spot CTRL2-2) being for the biological control of anti-Troponin I detection antibodies (monoclonal antibody 560, P2 binding partner) using a CTRL2 control ( epitope peptide coupled protein P2);
- CTRL2-1 and spot CTRL2-2 epitope peptide coupled protein P2
- FIG. 14 gives the signal results obtained in chambers 1 and 2 of the Magnotech cartridge according to FIG. 13 with three different samples, namely a negative TnI sample, and two TnI positive samples, having two different concentrations of TnI. ;
- FIG. 15 is a schematic representation of the biological control of the anti-Troponin I 19C7 monoclonal antibody (PI binding partner), using a CTRL1 control (protein coupled to the epitope peptide of PI and to a magnetic particle), in the part of a Magnotech technology used for the detection of troponin I as an analyte; and FIG. 16 gives the signal results obtained in a chamber exhibiting biological control of the PI binding partner according to FIG. 15, with two different samples, namely a negative TnI sample, and a positive TnI sample.
- CTRL1 control protein coupled to the epitope peptide of PI and to a magnetic particle
- Figure 1 is intended to illustrate the concept underlying the present invention. Said concept relates to increasing the detection sensitivity in the context of any type of sandwich-type immunoassay, through the significant reduction - or even the suppression - in the obtaining of false-negative results.
- a first PI binding partner in this case the antibody 10
- This antibody is used as a capture antibody.
- a second P2 binding partner in this case antibody 11, immobilizable by formation of a sandwich-type complex with analyte 12 and capture antibody 10.
- Antibody 11 is him, called “detection antibodies”.
- This detection antibody 11 is associated, directly or indirectly, with a detection element 111 making it possible to detect the formation of the "sandwich” between the capture antibody 10, the analyte 12 and the detection antibody 11; forming said sandwich revealing the presence of the desired analyte 12 within the test sample (unnumbered in Figure 1).
- the type of detection element 111 used depends on the detection means employed.
- the detection element 111 will consist, respectively, of a fluorogenic or chromogenic element (that is to say to obtain a fluorescent reaction or a colored reaction) , such as an enzyme in the presence of a substrate.
- a fluorogenic or chromogenic element that is to say to obtain a fluorescent reaction or a colored reaction
- the sandwich-type immunoassay is implemented via an opto-magnetic technology platform such as the "Magnotech” platform
- the detection element 111 can simply be constituted by a magnetic particle / ball. Indeed, this magnetic particle will influence the intensity of the reflected light signal at the level of the detection region and, thus, will give an indication of the formation of the "sandwich", thus revealing the presence / quantification or absence of the analyte 12 sought.
- the inventive concept on which the present invention is based requires the use of two positive controls:
- a first positive control 13 making it possible to test not only the presence but above all the functional character of the capture antibody 10.
- This positive control 13 comprises an epitope 131 (for example of peptide type) specifically recognized by the capture antibody 10 and a detection element 132, identical to or different from the detection element 111;
- control 14 making it possible to test not only the presence but above all the functional character of the detection antibody 11, said control 14 comprising an epitope 141 (for example of peptide type) specifically recognized by the detection antibody 11.
- the epitopes 131 and 141 may be identical, and therefore the antibodies 10 and 11 too, for example when the analyte has several identical epitopes.
- the 131 and 141 epitopes are different when the analyte has only different epitopes, as is the case, for example, for Troponin I.
- test result When there is no signal at the region of detection and / or quantification of the analyte and there is also a lack of signal at the level of the biological control region of the antibody of capture 16 and / or at the level of the biological control region of the detection antibody 17, the test result should be considered uninterpretable. In the absence of biological control of the capture antibody 10, at the level of the biological control region 16, the test result would have been wrongly considered as negative, while the analyte of interest was well and truly present. contained in the tested sample.
- the device according to the invention clearly makes it possible to identify the binding partner (s) (in this case the capture antibody 10 and / or the detection antibody 11) defective.
- the binding partner in this case the capture antibody 10 and / or the detection antibody 11
- This is particularly of interest in the production of kits comprising a support (plate, reaction cartridge, etc.) on which are immobilized (for example by adsorption) capture binding partners and a solution comprising the detection antibodies.
- reaction cartridge 2 used in an opto-magnetic immunoassay device of the "Magnotech” type is shown in FIG. 2.
- This reaction cartridge 2 comprises an inlet 21 intended to receive the liquid sample to be tested, a channel 22 allowing the flow (for example natural or by capillarity) of the sample to be tested to the reaction zone 23 comprising, in the example shown in FIG. 2, a single reaction chamber 231.
- the device according to the invention can be used for different types of immunoassay, and in particular in an opto-immunoassay.
- magnetic using the platform "Magnotech” developed by the company Philips (Eindhoven, the Netherlands), the principle is recalled below with reference to Figures 3A-3D.
- the capture and detection antibodies are maintained in dry form.
- the magnetic particles 33, coated with the detection antibodies 31 are resuspended in the presence of the liquid sample previously introduced at the inlet 21 of the reaction cartridge 2 shown in FIG. 2.
- the lower electromagnet 35 is activated in order to attract the magnetic particles 33 towards the detection region 36 on which are immobilized (for example by covalent bonding or adsorption) capture antibody 30, thus allowing, when the desired analyte 32 is present, the formation of a sandwich-type complex between the capture antibody 30, the analyte 32 and the detection antibody 31 carried by a particle magnetic 33.
- the lower electromagnet 35 is deactivated and the upper electromagnet 34 is activated in order to induce the removal of the unbound magnetic particles 33 at the level of the detection region 36 - and thus keeping only, at said detection region 36, the magnetic particles 33 at least one detection antibody 31 of which is bound to at least one capture antibody 30 via at least one desired analyte 32 (formation a sandwich-type complex between the three aforementioned entities), if the analyte of interest is present in the sample tested.
- the lower electromagnet 35 may be considered as a "connecting" electromagnet, while the upper electromagnet 34 may be considered as a "washing" electromagnet.
- the particles / magnetic beads present at the detection region 36 are detected using, as detection means 37, a 2-dimensional photographic sensor ("2D ", Such as a CCD sensor), the detection element being constituted by the magnetic particle 33.
- 2D 2-dimensional photographic sensor
- the detection step, represented in FIG. 3D is based on the principle of total frustrated total internal reflection (f-TIR), as described, for example, in Bruis et al. [1] or in the patent application WO2013 / 054230.
- the detection principle is as follows: an incident light beam 38 is emitted (for example by a light source 39 of the LED type) towards the detection region 36 with an incident angle greater than the critical angle (depending on the material constituting the reaction cartridge) so that the entirety of the incident light beam 38 is reflected into a reflected light beam 40 - referred to in this case as an "evanescent beam" - in the direction of the detection means 37.
- the detection principle is based on the following assumption: the intensity of the reflected light beam 40 is inversely proportional to the quantity of magnetic particles 33 present at the detection region 36. in other words, the decrease in the intensity of the reflected light beam 40 observed at the detection means 37 is proportional to the number of magnetic particles 33 linked to the level of the detection surface 36, and consequently to the concentration of the analyte 32 present in the sample tested.
- the device according to the invention is adapted to be implemented in an opto-magnetic immunoassay of "Magnotech" type.
- FIG. 4 A device according to a first embodiment of the first aspect of the invention is shown in FIG. 4 (top view).
- This device is a reaction cartridge similar to that shown in FIG. 2. It comprises an inlet 41, a channel 42 and a reaction zone 43, except that the reaction zone comprises two reaction chambers 431 and 432, which are not directly connected to the reaction zone 43. fluid communication.
- FIG. 5 schematically illustrates a first configuration of this first embodiment during the first step of the "Magnotech” operating protocol, as represented in FIG. 3A.
- the first reaction chamber 431 comprises a region of detection and / or quantification of the analyte 56 at which the antibody of capture 50.
- the formation of a sandwich-like complex between the detection antibody 51 (associated with the magnetic particle 53), the analyte 52 and the capture antibody 50 will be evidenced at said detection region 56 by attenuation of the reflected light signal 40 at the detection means 37, as shown schematically in FIG. 3D.
- the second reaction chamber 432 comprises a biological control region of the capture antibody 50, at which level, if the capture antibody 50 is functional, the formation of a complex between the 551 epitope of the biological control 55 (associated with the magnetic particle 53) and the capture antibody 50.
- the detection at the level of the biological control region of the capture antibody 57, by attenuation of the light beam 40 at the detection means 37, as shown diagrammatically in FIG. 3D.
- This second reaction chamber 432 also comprises a biological control region of the detection antibody 58, at which, if said detection antibody 51 is operational, a complex will form between said detection antibody 51 (bound to the particle Magnetic 53) and epitope 541 of biological control 54.
- a biological control region of the detection antibody 58 at which, if said detection antibody 51 is operational, a complex will form between said detection antibody 51 (bound to the particle Magnetic 53) and epitope 541 of biological control 54.
- the presence of this complex is demonstrated, at the level of the biological control region of the detection antibody 58, via attenuation of the reflected light beam 40 at the level of the detection means 37, as shown schematically in FIG.
- Figure 6 shows a second configuration of the first embodiment according to a first aspect of the invention.
- that shown in FIG. 6 comprises two reaction chambers 431 and 432.
- the reaction chamber 431 comprises both the detection region and / or quantification of the analyte 56 and the biological control region of the detection antibody 58.
- the reaction chamber 432 for its part, comprises the biological control region of the capture antibody 57.
- the region biological control of the capture antibody 57 in a reaction chamber separate from the chamber 431, comprising the detection and / or quantification region of the analyte 56 and the biological control region of the antibody 58, avoids a phenomenon of competition between the analyte 52 and the biological control 55 with respect to binding to the capture antibody 50 at the region of detection and / or quantification of the analyte 56.
- FIG. 7 is a top view of a device according to a second embodiment of the first aspect of the invention. More precisely, this FIG. 7 represents a device according to the invention, in the form of a reaction cartridge, that can be used in an opto-magnetic device of the "Magnotech" type.
- This reaction cartridge is similar to that shown in FIG. 4, except that the reaction zone 73 does not comprise two distinct reaction chambers but rather three distinct reaction chambers 731, 732 and 733, which are not in direct fluid communication.
- the first reaction chamber 731 comprises the detection and / or quantification region of the analyte 86, at which the complex between the capture antibody 80, the analyte 82 and the detection antibody 81 is formed. reserve that analyte 82 be present in the sample tested.
- the reaction chamber 732 comprises the biological control region of the detection antibody 88, at which the complex between the detection antibody 81, bound to the magnetic particle 83, is formed with the epitope 841 of the biological control 84 provided that the detection antibody 81 is functional.
- the third reaction chamber 733 comprises the biological control region of the capture antibody 87, at which a complex is formed between the 851 epitope, bound to the magnetic particle 83, of the biological control 85 and the capture antibody. 80, provided that the latter is operational / functional.
- the detection of the presence of these complexes at their respective detection zones 86, 88 and 87 is carried out as indicated above with reference to FIG. 3D.
- FIG. 9 A third embodiment according to a first aspect of the invention is shown diagrammatically in FIG. 9.
- the latter relates to a device according to the invention in the form of a reaction cartridge comprising a single reaction chamber, as shown in FIG.
- the single reaction chamber 99 comprises: a region for detecting and / or quantifying the analyte 96, at which a complex is formed between the capture antibody 90, the analyte 92 and the antibody detector 91 (associated with the magnetic particle 93) in the presence of the desired analyte 92;
- a biological control region of the detection antibody 98 at which a complex is formed between the detection antibody 91, bound to the magnetic particle 93, and the 941 epitope of the biological control 94, when the latter is functional;
- a biological control region of the capture antibody 97 at the level of which a complex is formed between the latter and the epitope 951, bound to the magnetic particle 93, of the biological control 95 (bound to a magnetic particle 93 ) when said capture antibody 90 is functional.
- the detection antibody 91 be present in excess with respect to its possible binding sites (consisting of analyte 92 and biological control 94) in order to avoid the competition phenomenon presented supra.
- capture antibody 90 it is also important that capture antibody 90 be present in excess of its possible binding sites (i.e. antigen 92 and 951 epitope of biological control 95).
- the epitope 951 of the biological control 95 in order to prevent the specific binding of the epitope 951 of the biological control 95 to the capture antibody 90, at the level of the detection and / or quantification region 96, preventing the formation of a complex of the type at the level of said detection and / or quantification region 96. It is also important that the epitope 951 of the biological control is not present in excess large quantity for the same reason. This is particularly important when the analyte is present in small amounts in the sample to be assayed.
- the epitope 951 of the biological control 95 binds not only to the capture antibody 90 at the biological control region of the capture antibody 97 but also at the region of detection and / or quantification of the analyte 96, it is important or even primordial that the magnetic particles 93 associated with the biological control 95 are associated, directly or indirectly with a marker 932 for detecting which complexes, at regions 96 and 97, are due to the binding of the biological control 95 to the capture antibody 90, and thereby indirectly infer which complexes are actually indicative of the presence of the analyte 92 in the test sample.
- the differentiation between different regions can also be performed using differently colored particles or particles of different diameter.
- the magnetic particles 93 associated with the detection antibodies 91 are labeled with a marker 931, in order to directly identify the sandwich-type complexes at the region of detection and / or quantification of the analyte 96.
- the magnetic particles 93 associated with the biological control 95 and the magnetic particles 93 associated with the detection antibodies 91 are respectively labeled with a marker 932 and 931, said markers 932 and 931 being different from one of the other.
- the present invention is applicable in various types of sandwich type immunoassay and in particular in rapid unit tests, also called tests / lateral flow assays.
- the patent application WO 2012/066235 in the name of the Applicant, discloses a device for performing a rapid unit test, which device incorporates a positive control. The operation of this device of the prior art is illustrated in FIGS. 10A-10D.
- the device according to WO 2012/066235 is a cassette comprising a support (not represented), a matrix 1001 comprising an application zone of the liquid sample 1002, a marking zone 1003, a viewing zone of the results 1005 a positive control zone 1006, an optional migration control zone 1007 and, optionally, an absorption zone of the sample 1008.
- Zone 1001 is shown as a rectangular strip whose longitudinal axis is in a horizontal position.
- Zones 1002, 1003, 1005, 1006, 1007 and 1008 are in fluid communication.
- Zone 1003 comprises a detectably labeled detection antibody (e.g., a colored latex particle, a gold particle, etc.).
- This labeled antibody can freely migrate through the 1001 template and react with the desired analyte (antigen in the present case) if it is present in the test sample.
- a capture antibody with a specificity for an antigen epitope different from that recognized by the detection antibody is immobilized (for example by covalent linkage or adsorption).
- an antigen analog is either immobilized directly or indirectly or can migrate freely in the presence of the fluid stream at the zone 1006 until it is immobilized by the capture antibody at the positive control zone 1006.
- Figures 10A, 10B and 10C summarize the operation of the test according to the patent application WO2012 / 066235.
- FIG. 10D comparable a priori to FIG. 10A, also illustrates the representation of a false negative result obtained using the aforementioned cassette as explained below.
- Figure 10B illustrates the result obtained with a positive control sample.
- the emission of a detectable signal 10051 ("test line") at the viewing area of the results 1005 is noted.
- This detectable signal 10051 is due to the formation of a complex sandwich type between the capture antibody, the antigen and the detection antibody, at the display area of the results 1005.
- the detectable signal 10051 strictly speaking, is due to the marker associated with the antibody of detection.
- test is such that it indicates that the sample comprises the analyte and is considered to be interpretable according to the patent application WO2012 / 066235.
- FIG. 10C illustrates a result regarded as "uninterpretable" in that no signal is detected either at the result viewing area 1005 or at the positive control region 1006.
- FIG. 10D By multiplying the tests with positive control samples (ie including the desired analyte), the Applicant surprisingly uncovered, in very small proportions, the profile shown in FIG. 10D, identical to that object of the present invention.
- Fig. 10A i.e., having no signal at the result display area 1005, but a signal 10061 at the positive control region 1006 and a signal 10071 at the migration control region 1007.
- FIGS. 11A, 11B, 11C, 11D, 11E and 11F illustrate the operation of a cassette similar to that described in WO2012 / 066235 but having an increased detection sensitivity.
- Figs. 11A-11F show top views of a lateral flow metering device according to a second aspect of the invention.
- this device comprises a support (not shown), a matrix 1101 (represented in the form of a rectangular strip whose longitudinal axis is in a horizontal position), said matrix comprising a zone of application of the sample 1102, a labeling zone 1103 and a reaction zone 1800.
- This reaction zone comprises a detection region of the analyte (in this case an antigen) 1105, a positive control region of the capture binding partner 1109 (for example, an antibody), a positive control binding region of the detection binding partner 1106 (e.g., an antibody) and, optionally, a migration control region 1107.
- the template 1101 may further optionally include an absorption zone of the sample 1108.
- the first analogue of the analyte CTRL1 (consisting for example of an epitope peptide of PI coupled to a marker or a precursor labeling) is disposed downstream (following the direction of migration of the liquid) of this detection region of the analyte 1105 but upstream of the positive control line of the capture antibody at which PI is immobilized, namely at the level of the region 1104 (located between the regions 1105 and 1109 ), or at a subregion of the positive control region of the capture antibody 1109, located upstream of said positive control line of the capture antibody, at which the antibody is immobilized.
- the positive control region of the detection antibody P2 1106, for its part, comprises the second analogue of the analyte CTRL2 immobilized at the level of the positive control line of the detection antibody P2.
- the marking area 1103 comprises the excess detection antibody P2.
- the positive control region of the capture partner PI 1109 where appropriate including the aforesaid subregion upstream of said positive control line of the capture antibody, as indicated above, can also be found after the control region. positive detection partner P2 1106 (not shown).
- the reaction zone 1800 then comprises, in order, a detection region of the analyte (in this case an antigen) 1105, a positive control region of the detection antibody 1106, a positive control region of the analyte. 1109 capture antibody and, optionally, a migration control region 1107.
- the template 1101 may further optionally include an absorption zone of the sample 1108.
- FIG. 11B illustrates the results obtained with the device of the invention after application of a so-called "negative control" sample, ie not containing the desired analyte. Since the sample is a negative control, there is, by definition, no emission of a detectable signal at the detection region of analyte 1105 (specifically at the test line). On the other hand, a detectable signal is emitted at the level of the positive control region of the capture antibody 1109 (signal 11091 at the positive control line of the capture antibody), at the level of the Positive control region of the detection antibody 1106 (signal 11061 at the positive control line of the detection antibody) and at the migration area 1107 (signal 11071 at the migration control line ), when the latter is present.
- a detectable signal is emitted at the level of the positive control region of the capture antibody 1109 (signal 11091 at the positive control line of the capture antibody), at the level of the Positive control region of the detection antibody 1106 (signal 11061 at the positive control line of the detection antibody) and at
- Figure 11C illustrates the results obtained with the device of the invention after applying a positive sample for a predetermined analyte. As shown in FIG.
- the sample being positive for the predetermined analyte (antigen)
- there is emission of a detectable signal at the detection region of analyte 1105 (signal 11051 at the line test), at the level of the positive control region of the capture antibody 1109 (11091 signal at the positive control line of the capture antibody) at the positive control region of the detection antibody 1106 (signal 11061 at the positive control line of the detection antibody ), as well as at the migration zone 1107 (signal 11071) when the latter is present.
- Figures 11D, 11E and 11F illustrate results obtained after applying a seemingly negative sample for an analyte to be determined.
- the presence of a signal 11091 at the positive control region of the capture antibody 1109 indicates that it is functional.
- the absence of a signal at the level of the positive control region of the detection antibody 1106 indicates that the functional character of this detection antibody is at least partially impaired. It can not be concluded that the sample is negative. We can only conclude that the test is uninterpretable.
- the device may comprise two segments S1 and S2, each comprising a zone for applying the sample.
- the first segment S1 comprises, besides the zone of application of the sample, a marking zone and a reaction zone comprising an analyte detection region and a positive control region of the detection link partner P2
- the second segment S2 comprises, in addition to the sample application area, the positive control region of the capture binding partner P1, the latter comprising a subregion at which the capture partner PI is immobilized (for example in the form of a positive control line of PI), the control CTRL1 being disposed upstream (in the direction of migration of the liquid) of said subregion as explained above.
- This embodiment is particularly interesting because the user can use this second segment S2 only if the reading of the first segment indicates prima facie the absence of analyte in the sample tested but the control CTRL2 partner detection link P2 is positive. In such a case, the user does not know if the result obtained is a true negative, namely that the desired analyte is absent from the tested sample, or if the capture binding partner PI is faulty. The user can easily decide between these two hypotheses thanks to the second segment S2 (for example by visual or optical reading of the latter). Moreover, this makes it possible to be able to replace the sample of the second segment S2 by an appropriate buffer interchangeable with the sample at this segment.
- Peptide synthesis Peptides containing the epitopes recognized by the anti-Troponin I 19C7 (SASRKLQLK) and 560 (ELTGLGFAELQ) antibodies (Hytest, Turku, Finland) were produced by chemical synthesis according to the procedures well known to those skilled in the art such as synthesis. solid phase peptide described by Merrifield, 1962 (Merrifield, 1962, J. Am Chem Soc 85: 2149) and Fields et al, 1990 (Fields GB, Noble RL, 1990, Int J Pept Protein Res., 35 (3): 161-214), using a polystyrene polymer containing 0.1-1.0 mMol amines / g of polymer.
- the peptides are deprotected and cleaved from the polymer in the presence of a mixture of trifluoroacetic acid-ethanedithiol-triisopropylsilane-water (94 / 2.5 / 1 / 2.5 V / V / V / V) for about 2 hours. .
- the peptides are extracted by precipitation in diethyl ether at 0 ° C. They are purified by techniques such as high performance liquid chromatography. Lyophilization of the appropriate purification fractions leads to a homogeneous peptide which is characterized by standard physicochemical techniques such as mass spectrometry, high performance liquid chromatography, amino acid analysis.
- Each epitope peptide is coupled to bovine serum albumin (BSA, Proliant, Ankeny, IA, USA) via sulfo-SMCC (25 mg / ml, Thermo Fischer Scientific Inc, Rockford, IL, USA).
- BSA bovine serum albumin
- sulfo-SMCC 25 mg / ml, Thermo Fischer Scientific Inc, Rockford, IL, USA. This chemical modification is performed at a ratio of 1: 10 BSA / SMCC during an incubation of 1 h at 30 ° C. +/- 1 ° C. and is followed by dialysis against a 50 mM PO 4 buffer, 150 mM NaCl pH 6.8.
- Each epitope peptide diluted to 5g / L in 50mM PO 4 buffer, 150mM NaCl, 5mM EDTA pH 6.8 is then contacted with BSA-SMMC in a ratio of 3 epitope peptide molecules per BSA molecule. After 18 h of incubation at 2-8 ° C, the coupling reaction is blocked by the addition of 2-aminoethanethiol (2MEA, Thermo Fisher Scientific Inc., Rockford, IL, USA) at a concentration respecting the equimolarity between the molecules. 2MEA and SMCC and incubation for 20 minutes with shaking. Each BSA-epitope peptide control is dialyzed against PBS buffer, 0.9g / L azide and stored at 2-8 ° C.
- EXAMPLE 2 Biological Control of the P2 Linking Partner with the CTRL2 Epitope Peptide
- the principle of this control in a test for the detection of troponin I as an analyte is shown in FIG. 12, PI 1200 and P2 1201 being antibodies anti-TnI monoclonals, respectively 19C7 and 560, P2 1201 being coupled to a magnetic particle 1203 as previously described and CTRL2 1204 being a P2 12041 epitope peptide coupled to BSA 12042, as previously described.
- the cartridge Magnotech (Philips, Eindhoven, Netherlands) consists of three elements that are: the optical part 1209, a sheet 1210 and a filtration unit on which is deposited the sample (not shown).
- the optical part is the lower part of the cartridge. It comprises the point of entry of the filtered sample, connected to two reaction chambers (chamber 1 and chamber 2) by a microfluidic channel.
- the sheet is a biocompatible adhesive film which constitutes the upper part of the cartridge and ensures the closure of the reaction chambers and fluidized channels etched in the optical part.
- the anti-Troponin I 19C7 antibody (PI binding partner) is deposited as a spot in each reaction chamber (spot PI 1300) by printing (sciFLEXARRAYER, SU, Scienion AG), as described in FIG. the article by Dittmer et al [7] (Journal of immunological methods, 338: 40-46), at a concentration of 40 ⁇ g / ml of antibody.
- Control CTRL2 1204 (BSA coupled to the epitope peptide of anti-Troponin I antibody 560), prepared in Example 1.2. and used as a biological control of the P2 binding partner (anti-troponin antibody ⁇ 560 coupled to particles Magnetic functionalized as described in Example 1.3. except that the antibody concentration is 60 ⁇ g of antibody per mg of magnetic particles), is also deposited in spot form at a concentration of 0.1 of CTRL2 (spot CRTL2-1 13041) or 1 ng / mL of CTRL2 (spot CRTL2-2 13042).
- the P2 bonding partner is deposited on the surface of the 1209 sheet of each reaction chamber using Nanodrop NS-2Stage (Innovadyne Technologies, Inc. Carnforth, UK). The three elements are then assembled and preserved in the presence of desiccant at
- the graphs in FIG. 14 show that: when the Troponin negative sample was assayed, a signal considered negative (less than 0.4%) was observed at the PI 1300 spots in both chambers, while the biological control of the P2 binding partner gave a positive signal (CTRL2-1 spot 13041 and CTRL2-2 spot 13042). This confirms that the 560 antibodies coupled to the beads are functional and that the sample is a true negative. In addition, it also shows that it is possible to perform the positive control indifferently in a chamber separate from the chamber where the analyte is detected or in the same chamber.
- troponin positive samples with concentrations of 1.05 ⁇ g / ml and 2.43 g / ml, respectively, show an increasing signal at the PI 1300 spots (signals from 14% to 41%). It is also observed that at the CTRL2 spots 13041 and 13042, no significant change between the signals obtained with the troponin I positive samples and those from the negative sample was detected. This also shows that troponin ⁇ present in the sample does not affect biological control because magnetic beads coupled with anti-troponin I 560 (P2) antibody are
- CTRL2 (CTRL2 spots 13041 and 13042), even at a high concentration of Troponin I.
- PI 1500 being an anti-TnI 19C7 monoclonal antibody
- CTRL1 1505 being an epitope peptide of PI 15051 coupled to the beads.
- magnetic 1503 as previously described.
- the cartridge is prepared as described in 2.1. above, except that the CTRLl 1505 control prepared in 1.3. above (magnetic ball 1503 coupled to the epitope peptide of the anti-troponin monoclonal antibody I 19C7 15051) is deposited on the sheet 1510 at the reaction chamber, opposite the binding partner PI 1500 (monoclonal antibody 19C7 fixed on the optical part (spot PI, not shown)).
- CTRLl 1505 control prepared in 1.3. above (magnetic ball 1503 coupled to the epitope peptide of the anti-troponin monoclonal antibody I 19C7 15051) is deposited on the sheet 1510 at the reaction chamber, opposite the binding partner PI 1500 (monoclonal antibody 19C7 fixed on the optical part (spot PI, not shown)).
- the graphs in FIG. 16 show that: when the Troponin I negative sample was assayed, a mean positive signal of 59% was obtained (spot CTRL1, not shown), which demonstrates that the binding partner PI 1500 ( 19C7 antibody) is functional;
Landscapes
- Health & Medical Sciences (AREA)
- Immunology (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Molecular Biology (AREA)
- Biomedical Technology (AREA)
- Chemical & Material Sciences (AREA)
- Hematology (AREA)
- Urology & Nephrology (AREA)
- Biotechnology (AREA)
- Microbiology (AREA)
- Cell Biology (AREA)
- Food Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Physics & Mathematics (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Investigating Or Analysing Biological Materials (AREA)
- Peptides Or Proteins (AREA)
Abstract
Description
Claims
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
FR1357754A FR3009386A1 (en) | 2013-08-02 | 2013-08-02 | DEVICE AND METHOD FOR BIOLOGICAL ANALYZES |
PCT/FR2014/052004 WO2015015130A1 (en) | 2013-08-02 | 2014-07-31 | Device and method for biological analyses |
Publications (1)
Publication Number | Publication Date |
---|---|
EP3028045A1 true EP3028045A1 (en) | 2016-06-08 |
Family
ID=49382440
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
EP14759031.9A Withdrawn EP3028045A1 (en) | 2013-08-02 | 2014-07-31 | Device and method for biological analyses |
Country Status (5)
Country | Link |
---|---|
US (1) | US20160169884A1 (en) |
EP (1) | EP3028045A1 (en) |
CN (1) | CN105705948B (en) |
FR (1) | FR3009386A1 (en) |
WO (1) | WO2015015130A1 (en) |
Families Citing this family (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US10267792B2 (en) * | 2016-09-09 | 2019-04-23 | International Business Machines Corporation | Device for detecting toxic shock syndrome toxins and method of making the same |
Family Cites Families (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US6686170B1 (en) * | 1994-08-17 | 2004-02-03 | Abbott Laboratories | Assay devices with mobile control reagents |
ES2322297T3 (en) * | 2006-01-14 | 2009-06-18 | F. Hoffmann-La Roche Ag | IMMUNOLOGICAL TEST ELEMENT WITH PERFECTED CONTROL AREA. |
US20100136531A1 (en) * | 2006-04-10 | 2010-06-03 | Tecra International Pty Ltd | Nucleic acid detection using lateral flow methods |
CN101473043A (en) * | 2006-04-10 | 2009-07-01 | 3M创新有限公司 | Nucleic acid detection using lateral flow methods |
FR2967497B1 (en) * | 2010-11-17 | 2014-12-12 | Biomerieux Sa | DEVICE AND METHOD FOR IMMUNOASSAY |
-
2013
- 2013-08-02 FR FR1357754A patent/FR3009386A1/en active Pending
-
2014
- 2014-07-31 EP EP14759031.9A patent/EP3028045A1/en not_active Withdrawn
- 2014-07-31 WO PCT/FR2014/052004 patent/WO2015015130A1/en active Application Filing
- 2014-07-31 US US14/909,697 patent/US20160169884A1/en not_active Abandoned
- 2014-07-31 CN CN201480043623.5A patent/CN105705948B/en not_active Expired - Fee Related
Non-Patent Citations (2)
Title |
---|
None * |
See also references of WO2015015130A1 * |
Also Published As
Publication number | Publication date |
---|---|
US20160169884A1 (en) | 2016-06-16 |
CN105705948B (en) | 2018-02-06 |
WO2015015130A1 (en) | 2015-02-05 |
FR3009386A1 (en) | 2015-02-06 |
CN105705948A (en) | 2016-06-22 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
EP1917529B1 (en) | Analyte assaying by means of immunochromatography with lateral migration | |
Dutra et al. | Surface plasmon resonance immunosensor for human cardiac troponin T based on self-assembled monolayer | |
KR102302678B1 (en) | Method of agglutination immunoassay | |
Frasconi et al. | Surface plasmon resonance immunosensor for cortisol and cortisone determination | |
FR2997194A1 (en) | DEVICE FOR DETERMINING AT LEAST ONE ANALYTE LIKELY TO BE CONTAINED IN A LIQUID SAMPLE | |
JPH06501555A (en) | Improvements in solid-phase binding assays | |
FR2881828A1 (en) | Quantitative measurement of analytes in a blood plasma sample by immunochromatography, comprises contacting the sample with control reagent, depositing the sample on application zone of support and measuring intensity of test signal | |
EP2641088B1 (en) | Device and method for immunoassays | |
JP6810055B2 (en) | How to reuse test probes and reagents in immunoassay | |
JP2012242162A (en) | Composite particle as indicator | |
EP1040353B1 (en) | Method for detecting an analyte and kit | |
WO2012172232A1 (en) | Device and method for immunoassays | |
EP1608978B1 (en) | Solid-phase immunochromatographic methods | |
US20220317128A1 (en) | Method for detecting particulate substance using immunochromatography, and kit for same | |
EP3028045A1 (en) | Device and method for biological analyses | |
WO2009115756A2 (en) | Method for direct detection of ischaemia-modified albumin using a partner for binding to an aldehyde derivative resulting from the peroxidation of lipids in bound form | |
US20100285607A1 (en) | Method for detecting and quantifying analytes on a solid support with liposome-encapsulated fluorescent molecules | |
FR3067814A1 (en) | METHOD FOR APPLYING, ON A SOLID SUPPORT, AT LEAST ONE MOLECULAR BINDING PARTNER | |
FR3113318A1 (en) | System for the rapid analysis of a biological sample, intended for the detection of the presence of at least one analyte in said biological sample | |
KR101594379B1 (en) | Method for Detection of Protein Comprising Histidine-Tag Using immunochromatography | |
Mao et al. | Detection of autoantibodies for type 1 diabetes using label-free optical sensors | |
EP1844332A1 (en) | Immunochromatographic method for the quantitative measurement of analytes in a liquid sample | |
WO2021255021A1 (en) | System for rapid analysis of a biological sample for distinguishing specific serological antibodies of an infectious agent present in the biological sample | |
FR3111431A1 (en) | System for the rapid analysis of a biological sample, intended for the detection of the presence of at least one analyte in said biological sample |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
PUAI | Public reference made under article 153(3) epc to a published international application that has entered the european phase |
Free format text: ORIGINAL CODE: 0009012 |
|
17P | Request for examination filed |
Effective date: 20160215 |
|
AK | Designated contracting states |
Kind code of ref document: A1 Designated state(s): AL AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HR HU IE IS IT LI LT LU LV MC MK MT NL NO PL PT RO RS SE SI SK SM TR |
|
AX | Request for extension of the european patent |
Extension state: BA ME |
|
RIN1 | Information on inventor provided before grant (corrected) |
Inventor name: RENARD, NATHALIE |
|
DAX | Request for extension of the european patent (deleted) | ||
STAA | Information on the status of an ep patent application or granted ep patent |
Free format text: STATUS: EXAMINATION IS IN PROGRESS |
|
17Q | First examination report despatched |
Effective date: 20170309 |
|
GRAP | Despatch of communication of intention to grant a patent |
Free format text: ORIGINAL CODE: EPIDOSNIGR1 |
|
STAA | Information on the status of an ep patent application or granted ep patent |
Free format text: STATUS: GRANT OF PATENT IS INTENDED |
|
INTG | Intention to grant announced |
Effective date: 20171110 |
|
STAA | Information on the status of an ep patent application or granted ep patent |
Free format text: STATUS: THE APPLICATION IS DEEMED TO BE WITHDRAWN |
|
18D | Application deemed to be withdrawn |
Effective date: 20180321 |