Human thiopurine S-methyltransferase (TPMT) ELISA kit

Code CSB-E17858h
Size 96T,5×96T,10×96T
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Product Details

Target Name
thiopurine S-methyltransferase
Alternative Names
HGNC:12014 ELISA Kit; S adenosyl L methionine thiopurine S methyltransferase ELISA Kit; Thiopurine methyltransferase ELISA Kit; Thiopurine S methyltransferase ELISA Kit; Thiopurine S-methyltransferase ELISA Kit; TPMT ELISA Kit; TPMT_HUMAN ELISA Kit
Abbreviation
TPMT
Uniprot No.
Species
Homo sapiens (Human)
Sample Types
serum, plasma, tissue homogenates
Detection Range
1.56 mU/mL-100 mU/mL
Sensitivity
0.39 mU/mL
Assay Time
1-5h
Sample Volume
50-100ul
Detection Wavelength
450 nm
Research Area
Signal Transduction
Assay Principle
quantitative
Measurement
Sandwich
Precision
Intra-assay Precision (Precision within an assay): CV%<8%      
Three samples of known concentration were tested twenty times on one plate to assess.  
Inter-assay Precision (Precision between assays): CV%<10%      
Three samples of known concentration were tested in twenty assays to assess.    
             
Linearity
To assess the linearity of the assay, samples were spiked with high concentrations of human TPMT in various matrices and diluted with the Sample Diluent to produce samples with values within the dynamic range of the assay.
  Sample Serum(n=4)  
1:200 Average % 87  
Range % 80-92  
1:400 Average % 98  
Range % 90-105  
1:800 Average % 100  
Range % 91-110  
1:1600 Average % 93  
Range % 86-98  
Recovery
The recovery of human TPMT spiked to levels throughout the range of the assay in various matrices was evaluated. Samples were diluted prior to assay as directed in the Sample Preparation section.
Sample Type Average % Recovery Range  
Serum (n=5) 93 85-100  
EDTA plasma (n=4) 92 80-100  
             
             
Typical Data
These standard curves are provided for demonstration only. A standard curve should be generated for each set of samples assayed.
mU/ml OD1 OD2 Average Corrected  
100 2.239 2.152 2.196 2.044  
50 1.854 1.760 1.807 1.655  
25 1.311 1.294 1.303 1.151  
12.5 0.843 0.839 0.841 0.689  
6.25 0.571 0.534 0.553 0.401  
3.12 0.380 0.359 0.370 0.218  
1.56 0.243 0.239 0.241 0.089  
0 0.153 0.151 0.152    
Materials provided
  • A micro ELISA plate --- The 96-well plate has been pre-coated with an anti-human TPMT antibody. This dismountable microplate can be divided into 12 x 8 strip plates.
  • Two vials lyophilized standard ---Dilute a bottle of the standard at dilution series, read the OD values, and then draw a standard curve.
  • One vial Biotin-labeled TPMT antibody (100 x concentrate) (120 μl/bottle) ---Act as the detection antibody.
  • One vial HRP-avidin (100 x concentrate) (120 μl/bottle) ---Bind to the detection antibody and react with the TMB substrate to make the solution chromogenic.
  • One vial Biotin-antibodyDiluent (15 ml/bottle) ---Dilute the Biotin-antibody.
  • One vial HRP-avidin Diluent (15 ml/bottle) ---Dilute the HRP-avidin solution.
  • One vial Sample Diluent (50 ml/bottle)---Dilute the sample to an appropriate concentration.
  • One vial Wash Buffer (25 x concentrate) (20 ml/bottle) ---Wash away unbound or free substances.
  • One vial TMB Substrate (10 ml/bottle) ---Act as the chromogenic agent. TMB interacts with HRP, eliciting the solution turns blue.
  • One vial Stop Solution (10 ml/bottle) ---Stop the color reaction. The solution color immediately turns from blue to yellow.
  • Four Adhesive Strips (For 96 wells) --- Cover the microplate when incubation.
  • An instruction manual
Materials not provided
  • A microplate reader capable of measuring absorbance at 450 nm, with the correction wavelength set at 540 nm or 570 nm.
  • An incubator can provide stable incubation conditions up to 37°C±5°C.
  • Centrifuge
  • Vortex
  • Squirt bottle, manifold dispenser, or automated microplate washer
  • Absorbent paper for blotting the microtiter plate
  • 50-300ul multi-channel micropipette
  • Pipette tips
  • Single-channel micropipette with different ranges
  • 100ml and 500ml graduated cylinders
  • Deionized or distilled water
  • Timer
  • Test tubes for dilution
Troubleshooting
and FAQs
Storage
Store at 2-8°C. Please refer to protocol.
Lead Time
3-5 working days after you place the order, and it takes another 3-5 days for delivery via DHL or FedEx
Description

This Human TPMT ELISA Kit was designed for the quantitative measurement of Human TPMT protein in serum, plasma, tissue homogenates. It is a Sandwich ELISA kit, its detection range is 1.56 mU/mL-100 mU/mL and the sensitivity is 0.39 mU/mL.

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Target Background

Function
(From Uniprot)
Catalyzes the S-methylation of thiopurine drugs such as 6-mercaptopurine (also called mercaptopurine, 6-MP or its brand name Purinethol) and 6-thioguanine (also called tioguanine or 6-TG) using S-adenosyl-L-methionine as the methyl donor. TPMT activity modulates the cytotoxic effects of thiopurine prodrugs. A natural substrate for this enzyme has yet to be identified.
Gene References into Functions
  1. Mutation rate of NUDT15 in Chinese inflammatory bowel disease patients is higher than that of TPMT. NUDT15 polymorphism is a better predictor for AZA-induced leukopenia than TPMT polymorphism. PMID: 29491687
  2. In Mexican patients, mutant alleles were detected in 6.2 and 5.2% of systemic lupus erythematosus and rheumatoid arthritis cases, respectively. PMID: 29264794
  3. TPMT*3C affects TPMT activity in Chinese patients with neuromyelitis optica spectrum disorders. PMID: 29191122
  4. Patients also developed infection who mostly (71%) needed hospitalization. None of the studied G238C, G460A and A719G TPMT variant alleles were detected. Infections and febrile neutropenia were common causes of 6-PM dose modification and interruption. PMID: 29321348
  5. ANCA associated vasculitis (AAV) patients treated with cyclophosphamide induction and azathioprine maintenance therapy were included and followed for 60 months. TPMT genotype and activity were not independent predictors of relapse, and could not predict leukopenia or other adverse effects from azathioprine. PMID: 29630648
  6. Results show no association between TPMT or COMT single-nucleotide polymorphisms and cisplatin-induced ototoxicity. PMID: 28445188
  7. Of the 14 patients intolerant to thiopurines, NGS identified deleterious TPMT variants in 5 individuals whereas the biochemical test identified 8 individuals as intolerant (sensitivity 35.7% and 57.14%; specificity 93.75% and 50% respectively). SKAT-O test identified a significant association between MOCOS gene and TPMT activity (p = 0.0015), not previously reported. PMID: 27703193
  8. A statistically significant correlation between the presence of the G460A mutation and the development of leucopenia after the administration of thiopurines was observed. PMID: 28857898
  9. genetic association studies in cohort in Denmark: Data suggest that standard immunosuppression treatment failure in patients with autoimmune hepatitis is not associated with thiopurine S-methyltransferase genetic variants or HLA-DRB1*03 genotype in the population studied. (HLA-DRB1 = major histocompatibility complex class II DR beta 1) PMID: 28374975
  10. This article reviews the thiopurine pharmacogenetics and the methods applied in common practice to evaluate patient's TPMT status. [review] PMID: 28552060
  11. study aimed to determine the relationship between thiopurine metabolite levels and therapeutic response, and to investigate the association of NUDT15, TPMT, and thiopurine metabolites with leukopenia in patients with CD PMID: 29206869
  12. TPMT*3C is overrepresented in acute lymphoblastic leukemia (ALL) cases in comparison with non-ALL group PMID: 28476189
  13. TMPT polymorphisms are associated with 6-mercaptopurine intolerance in children treated for acute lymphoblastic leukemia. PMID: 27577869
  14. The frequency of TPMT mutation in a homogeneous Sicilian cohort of patients with inflammatory bowel disease, autoimmune and hematological disorders was low. TPMT genotyping is not a sensitive tool for predicting thiopurine-induced leukopenia. PMID: 27665263
  15. meta-analysis of three independent GWAS of TPMT activity; only genetic variants mapping to chromosome 6, including the TPMT gene region, were significantly associated with TPMT activity;finding is consistent with TPMT genotype being the primary determinant of TPMT activity PMID: 27770449
  16. In a GWAS, TPMT activity in patients behaves as a monogenic trait, further bolstering the utility of TPMT genetic testing in the clinic PMID: 27564568
  17. Review/Meta-analysis: thiopurine S-methyltransferase testing for averting drug toxicity. PMID: 27217052
  18. A point mutation in the thiopurine S-methyltransferase gene that led to exon 5 deletion in the transcribed mRNA. PMID: 26974142
  19. The aims of this research were to investigate the association of TPMT phenotypes with MDR1 genotypes. PMID: 27453281
  20. The aim of the study was to investigate frequencies of TPMT and ITPA polymorphisms in Lithuanian inflammatory bowel disease patients and analyze their association with azathioprine-related adverse events. PMID: 26674571
  21. With respect to TPMT, the variants TPMT*2 and TPMT*3A were not implicated in genetic susceptibility to childhood acute lymphoblastic leukemia PMID: 26845729
  22. structure-function relationships of TPMT PMID: 26410243
  23. The TPMT promoter region may serve as a pharmacogenomic biomarker when introducing thiopurine therapy PMID: 26411491
  24. The most frequently occurring nonfunctional TPMT allele in Croatian population is TPMT*3A. Variant genotypes were statistically significantly more common in Crohn's disease subgroup than in ulcerative colitis subgroup. PMID: 27333713
  25. Association of TPMT polymorphisms with overall azathioprine-induced adverse drug reactions, bone marrow toxicity and gastric intolerance, but not with hepatotoxicity (meta-analysis). PMID: 26633017
  26. association of TPMT polymorphisms with overall thiopurine-induced adverse drug reactions PMID: 25799415
  27. Identification of TPMT variants and subsequent dose reduction reduces hematologic events during thiopurine treatment of inflammatory bowel disease. PMID: 26072396
  28. TPMT*3c mutant allele was associated with azathioprine side effects (leukopenia, alopecia) in Chinese systemic lupus erythematosus patients. PMID: 24322830
  29. refinements in risk stratification and treatment have reduced the influence of TPMT genotype on treatment outcome in a contemporary protocol. PMID: 25940902
  30. We report on the presence of the TPMT*3C and *3A mutant alleles in the Libyan population. Therefore, monitoring the patients to be treated with doses of thiopurine drugs for TPMT variants is worthwhile to avoid the development of severe myelosuppression. PMID: 25819542
  31. this paper shows that the influence of TPMT and COMT on the development of cisplatin-induced hearing loss may be less important than previously suggested. PMID: 25551397
  32. Results show complete sequence-based screening study evaluating all TPMT variants in Asian populations some of them may ne of relevance in Korean population. PMID: 25564374
  33. distribution of the most frequent variants of TPMT gene was similar in a healthy population and patients with inflammatory bowel disease PMID: 23581716
  34. The prevalence of TPMT genotypes was high among Brazilian patients. Variants genes 2 and 3C may be associated with azathioprine pancreatic toxicity in a IBD southeastern Brazilian population PMID: 24696613
  35. Data suggest risk of myelotoxicity of high-dose methotrexate during methotrexate/6-mercaptopurine maintenance chemotherapy of childhood acute leukemia is not associated with heterozygosity in thiopurine methyltransferase in Nordic population studied. PMID: 25347948
  36. TMPT genotyping appears an important tool to further optimize 6-MP treatment design in Chilean patients with ALL PMID: 24774509
  37. suggest that germline genetic variation in TPMT and MTHFR do not significantly alter SOS risk in patients exposed to thioguanine PMID: 24737678
  38. TPMT*37 introduces a premature stop codon at position 216, resulting in loss of the last 29 amino acid residues from the C terminal of the TPMT protein. PMID: 24710034
  39. plays a key role in the metabolism of azathioprine/6- mercaptopurine (6-MP). Mutations in the enzyme lead to generation of excess thioguanine, which causes neutropenia via suppression of neutrophils. PMID: 23996738
  40. Activity measurement performed at diagnosis provides clinicians with information on immediate pharmacokinetic-related adverse events and/or hypermetabolism PMID: 24643197
  41. Heart transplantation recipients with TPMT genetic variant alleles who are treated with AZA develop acute rejection earlier, more frequently, and of greater severity. PMID: 24121523
  42. The study focused on explaining how a locally occurred TPMT A167G substitution propagated through hydrogen bonds alteration to induce structural modification which affects both thiopurine and S-adenosylmethionine receptors. PMID: 23025308
  43. This study shows the prevalence of TPMT genetic polymorphisms in population of Slovak IBD patients PMID: 23731044
  44. TPMT*3A and the TPMT*3C, which cause the largest decrease in enzyme activity, were both variant alleles detected in the Tunisian population PMID: 23553048
  45. In childhood acute lymphoblastic leukaemia, TPMT activity should not be used to predict heterozygosity particularly in blood samples obtained at disease diagnosis PMID: 23252716
  46. differential contribution of the enzyme TPMT to the cytotoxicity of the two thiopurines is probably due to its role in formation of the meTIMP, the cytotoxic methylated metabolite of 6-MP PMID: 23811272
  47. A predictive model combining variants in TPMT, ABCC3, and COMT with clinical variables significantly improved the prediction of hearing-loss development as compared with using clinical risk factors alone PMID: 23588304
  48. our results indicated that TPMT or COMT genetic variation was not related to cisplatin ototoxicity in children with cancer and did not influence cisplatin-induced hearing damage in laboratory models PMID: 23820299
  49. A high frequency and diversity of variant TPMT genotypes was found in this series with predominance of the TPMT*3B allele PMID: 23844534
  50. In a study of Italian myasthenia gravis patients treated with azathioprine, a new TPMT haplotype, TPMT* 3E, was observed only in association with intolerance. PMID: 23400745

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Subcellular Location
Cytoplasm.
Protein Families
Class I-like SAM-binding methyltransferase superfamily, TPMT family
Database Links

HGNC: 12014

OMIM: 187680

KEGG: hsa:7172

STRING: 9606.ENSP00000312304

UniGene: Hs.444319

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