EP2612145A2 - Dosage cellulaire destiné à mesurer l'activité biologique d'anticorps auto-immuns anti-tshr - Google Patents
Dosage cellulaire destiné à mesurer l'activité biologique d'anticorps auto-immuns anti-tshrInfo
- Publication number
- EP2612145A2 EP2612145A2 EP11751888.6A EP11751888A EP2612145A2 EP 2612145 A2 EP2612145 A2 EP 2612145A2 EP 11751888 A EP11751888 A EP 11751888A EP 2612145 A2 EP2612145 A2 EP 2612145A2
- Authority
- EP
- European Patent Office
- Prior art keywords
- tshr
- cells
- bioassay
- signal
- sample
- 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- Withdrawn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
- 230000001363 autoimmune Effects 0.000 title claims abstract description 37
- 230000004071 biological effect Effects 0.000 title claims abstract description 13
- 101150098159 TSHR gene Proteins 0.000 title claims description 3
- 238000005259 measurement Methods 0.000 title description 14
- 238000000423 cell based assay Methods 0.000 title description 2
- 102000003911 Thyrotropin Receptors Human genes 0.000 claims abstract description 105
- 108090000253 Thyrotropin Receptors Proteins 0.000 claims abstract description 105
- 238000004166 bioassay Methods 0.000 claims abstract description 52
- 239000013610 patient sample Substances 0.000 claims abstract description 42
- 239000000523 sample Substances 0.000 claims abstract description 40
- 230000004936 stimulating effect Effects 0.000 claims abstract description 35
- 230000000903 blocking effect Effects 0.000 claims abstract description 31
- 230000000638 stimulation Effects 0.000 claims abstract description 24
- 239000013074 reference sample Substances 0.000 claims abstract description 22
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- 108090000765 processed proteins & peptides Proteins 0.000 claims description 14
- 229920001184 polypeptide Polymers 0.000 claims description 13
- 102000004196 processed proteins & peptides Human genes 0.000 claims description 13
- 102000002260 Alkaline Phosphatase Human genes 0.000 claims description 6
- 108020004774 Alkaline Phosphatase Proteins 0.000 claims description 6
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- 241000699802 Cricetulus griseus Species 0.000 claims description 2
- 210000001672 ovary Anatomy 0.000 claims description 2
- 102000011923 Thyrotropin Human genes 0.000 claims 4
- 108010061174 Thyrotropin Proteins 0.000 claims 4
- 239000004017 serum-free culture medium Substances 0.000 claims 1
- 238000003556 assay Methods 0.000 description 33
- 210000002966 serum Anatomy 0.000 description 18
- 208000015023 Graves' disease Diseases 0.000 description 16
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- 238000013519 translation Methods 0.000 description 3
- 238000008940 Alkaline Phosphatase assay kit Methods 0.000 description 2
- TWRXJAOTZQYOKJ-UHFFFAOYSA-L Magnesium chloride Chemical compound [Mg+2].[Cl-].[Cl-] TWRXJAOTZQYOKJ-UHFFFAOYSA-L 0.000 description 2
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- XZKIHKMTEMTJQX-UHFFFAOYSA-N 4-Nitrophenyl Phosphate Chemical compound OP(O)(=O)OC1=CC=C([N+]([O-])=O)C=C1 XZKIHKMTEMTJQX-UHFFFAOYSA-N 0.000 description 1
- 102100024321 Alkaline phosphatase, placental type Human genes 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 1
- 241000581444 Clinidae Species 0.000 description 1
- 108020004414 DNA Proteins 0.000 description 1
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- 102000003688 G-Protein-Coupled Receptors Human genes 0.000 description 1
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- 101000772267 Homo sapiens Thyrotropin receptor Proteins 0.000 description 1
- 241001135569 Human adenovirus 5 Species 0.000 description 1
- 229930182555 Penicillin Natural products 0.000 description 1
- JGSARLDLIJGVTE-MBNYWOFBSA-N Penicillin G Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 JGSARLDLIJGVTE-MBNYWOFBSA-N 0.000 description 1
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- 239000012894 fetal calf serum Substances 0.000 description 1
- 230000001295 genetical effect Effects 0.000 description 1
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- 229940088597 hormone Drugs 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 210000005260 human cell Anatomy 0.000 description 1
- 208000003532 hypothyroidism Diseases 0.000 description 1
- 230000002989 hypothyroidism Effects 0.000 description 1
- 210000000987 immune system Anatomy 0.000 description 1
- 238000000099 in vitro assay Methods 0.000 description 1
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- 230000004962 physiological condition Effects 0.000 description 1
- 230000001817 pituitary effect Effects 0.000 description 1
- 108010031345 placental alkaline phosphatase Proteins 0.000 description 1
- 239000013612 plasmid Substances 0.000 description 1
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- 108020003175 receptors Proteins 0.000 description 1
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- 230000010473 stable expression Effects 0.000 description 1
- 229960005322 streptomycin Drugs 0.000 description 1
- 238000001890 transfection Methods 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
- G01N33/74—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving hormones or other non-cytokine intercellular protein regulatory factors such as growth factors, including receptors to hormones and growth factors
- G01N33/76—Human chorionic gonadotropin including luteinising hormone, follicle stimulating hormone, thyroid stimulating hormone or their receptors
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/90—Enzymes; Proenzymes
- G01N2333/914—Hydrolases (3)
- G01N2333/916—Hydrolases (3) acting on ester bonds (3.1), e.g. phosphatases (3.1.3), phospholipases C or phospholipases D (3.1.4)
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2500/00—Screening for compounds of potential therapeutic value
- G01N2500/10—Screening for compounds of potential therapeutic value involving cells
Definitions
- the problem underlying the invention is the provision of a precise, easy-to-perform and cost-efficient method to charac ⁇ terize and quantify the biological activity of anti TSHR autoimmune antibodies in patient samples .
- Fig. 3 shows the reproducibility of the bioassay.
- a "patient sample” according to the invention comprises at least blood serum or plasma from a human who may have Graves' disease (hyperthyroidism) or hypothyroidism.
- the SU values measured in all sample / cell line lot combinations are shown as data in the diagram of Fig. 2.
- the results demonstrate an intra-assay variance and an inter-assay variability below 10% over the whole range of measured SU values. This means that the assay produces accurate reproducible SU values for patient sera (samples B, C) and plasma (sample D) .
- Sample E further shows that even at higher concentrations of TSHR stimulating agent (in. this case TSH) , the assay leads to highly reproducible results .
- Example 5 Comparison of the clinical score of patients with the SU values of samples from the same patients determined by the bioassay according to the invention.
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Molecular Biology (AREA)
- Engineering & Computer Science (AREA)
- Immunology (AREA)
- Chemical & Material Sciences (AREA)
- Biomedical Technology (AREA)
- Urology & Nephrology (AREA)
- Endocrinology (AREA)
- Hematology (AREA)
- Physics & Mathematics (AREA)
- Microbiology (AREA)
- Biotechnology (AREA)
- Reproductive Health (AREA)
- Food Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Cell Biology (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
La présente invention concerne un dosage biologique destiné à déterminer l'activité biologique de stimulation d'anticorps auto-immuns dirigés contre le récepteur TSHR comprenant les étapes consistant à a) faire incuber des cellules exprimant TSHR avec un échantillon patient, b) détecter l'intensité d'un signal dans le surnageant des cellules, ledit signal étant entraîné par une modification de la concentration en AMPc dans les cellules en raison de la stimulation de TSHR, c) déterminer l'activité de stimulation d'anticorps auto-immuns anti-TSHR par comparaison de l'intensité du signal de l'étape b) à l'intensité du signal obtenu après l'incubation d'un premier échantillon de référence avec des cellules exprimant TSHR ; l'invention concerne également un dosage biologique destiné à déterminer l'activité biologique d'anticorps auto-immuns bloquants dirigés contre le récepteur de la thyréostimuline (TSHR), qui comprend les étapes consistant à a) faire incuber des cellules exprimant TSHR avec un échantillon patient, b) détecter l'intensité d'un signal dans le surnageant des cellules, ledit signal étant entraîné par une modification de la concentration en AMPc dans les cellules en raison de la stimulation de TSHR, c) déterminer l'activité de stimulation/blocage d'auto-anticorps anti-TSHR par comparaison de l'intensité du signal de l'étape b) aux intensités des signaux obtenus après l'incubation de premier et second échantillons de référence avec des cellules exprimant TSHR ; l'échantillon d'incubation de l'étape a) et le second échantillon de référence contenant la même quantité d'agent stimulant TSHR.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| EP11751888.6A EP2612145A2 (fr) | 2010-08-31 | 2011-08-31 | Dosage cellulaire destiné à mesurer l'activité biologique d'anticorps auto-immuns anti-tshr |
Applications Claiming Priority (3)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| EP10009042 | 2010-08-31 | ||
| EP11751888.6A EP2612145A2 (fr) | 2010-08-31 | 2011-08-31 | Dosage cellulaire destiné à mesurer l'activité biologique d'anticorps auto-immuns anti-tshr |
| PCT/EP2011/065035 WO2012028654A2 (fr) | 2010-08-31 | 2011-08-31 | Dosage cellulaire destiné à mesurer l'activité biologique d'anticorps auto-immuns anti-tshr |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| EP2612145A2 true EP2612145A2 (fr) | 2013-07-10 |
Family
ID=44545728
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| EP11751888.6A Withdrawn EP2612145A2 (fr) | 2010-08-31 | 2011-08-31 | Dosage cellulaire destiné à mesurer l'activité biologique d'anticorps auto-immuns anti-tshr |
Country Status (2)
| Country | Link |
|---|---|
| EP (1) | EP2612145A2 (fr) |
| WO (1) | WO2012028654A2 (fr) |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107271680B (zh) * | 2017-05-22 | 2019-04-23 | 中山大学附属第三医院 | 人髓鞘少突胶质细胞糖蛋白自身免疫抗体的检测方法 |
| WO2020050208A1 (fr) * | 2018-09-05 | 2020-03-12 | ヤマサ醤油株式会社 | Procédé et kit de mesure rapide de l'activité d'un auto-anticorps par rapport au récepteur tsh |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0327960A1 (fr) | 1988-02-11 | 1989-08-16 | F. Hoffmann-La Roche Ag | Formes sécrétables de phosphatase alcaline |
| GB9720693D0 (en) * | 1997-09-30 | 1997-11-26 | Univ Wales Medicine | Assay |
-
2011
- 2011-08-31 EP EP11751888.6A patent/EP2612145A2/fr not_active Withdrawn
- 2011-08-31 WO PCT/EP2011/065035 patent/WO2012028654A2/fr not_active Ceased
Non-Patent Citations (1)
| Title |
|---|
| See references of WO2012028654A2 * |
Also Published As
| Publication number | Publication date |
|---|---|
| WO2012028654A2 (fr) | 2012-03-08 |
| WO2012028654A3 (fr) | 2015-07-02 |
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