ADAMTS7 BioAssay™ ELISA Kit (Human)

ADAMTS (a disintegrin and metalloproteinase domain with thrombospondin type-1 modules) is a family of zinc-dependent proteases that are implicated in a variety of normal and pathological conditions, including arthritis and cancer. ADAMTS protein family members contain an N-terminal propeptide domain, a metalloproteinase domain, a disintegrin-like domain and a C-terminus that contains a varying number of thrombospondin type-1 (TSP-1) motifs. ADAMTS genes are primarily expressed in fetal tissues, including lung, kidney and liver. ADAMTS7 (ADAM metallopeptidase with thrombospondin type 1 motif, 7), also known as COMPase, is a 1686aa protein that exists as two alternatively spliced isoforms. Encoded by a gene that maps to human chromosome 15q25.1, ADAMTS7 contains eight TSP-1 motifs and binds one zinc ion per subunit. ADAMTS-7 is expressed in heart, brain, placenta, lung, liver, skeletal muscle, kidney and pancreas. ADAMTS7 is also located in meniscus, bone, tendon, cartilage, synovium, fat and ligaments, and is up-regulated in articular cartilage and synovium in arthritis patients. ADAMTS7 functions as a metalloprotease and may play a role in the degradation of COMP. ADAMTS7 is pH dependent, with optimum pH between 7.5 and 9.5.||ADAMTS7 (A disintegrin and metalloproteinase with thrombospondin Motifs 7) BioAssay™ ELISA Kit is a quantitative sandwich assay for the detection of ADAMTS7 in human serum, plasma and other biological fluids. Cell lysates and tissue homogenates, though not tested, may potentially be used as samples.||Detection Range:|0.313-20ng/ml||Sensitivity:|<0.188ng/ml||Precision: |Intra-Assay CV: <8%|Inter-Assay CV: <10%||Assay Principle:|This ELISA kit employs the sandwich enzyme-linked immunoassay technique, utilizing a microtiter plate pre-coated with an antibody specific to ADAMTS7. Standards and samples are added to the appropriate wells, then incubated. Aspirate. Do not wash the plate! The Antibody (Biotin) is added to all the wells. The plate is incubated and then washed. Streptavidin (HRP) is added to each microplate well and incubated then washed. Then the TMB substrate solution is added and incubated. After the TMB substrate solution is added, only those wells that contain ADAMTS7, the antibody (Biotin) and Streptavidin (HRP) will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulfuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450nm ± 10nm. The concentration of ADAMTS7 in the sample is then determined by comparing the O.D. of the sample to the standard curve.||Kit Components:|*353705A: Microtiter Strips, 1x96 wells (8x12 wells).|*353705B: Standard, 2x1 vial|353705C: Sample/Standard Dilution Buffer, 1x20ml|353705D: Antibody (Biotin) (Concentrated), 1x120ul|353705E: Antibody Dilution Buffer, 1x10ml|353705F: Streptavidin (HRP) (SABC), 1x120ul|353705G: SABC Dilution Buffer, 1x10ml|353705H: TMB Substrate, 1x10ml|353705J: Stop Solution, 1x10ml|353705K: Wash Buffer, 25X, 1x30ml||Storage and Stability:|Store unopened *353705A at 4ºC; store at -20ºC once opened. Store unopened *353705B at 4°C; once reconstituted, store at 4°C for up to 12 hours or at -20°C for up to 48 hours. Store other components at 4°C. Kit is stable for 6 months after receipt. For maximum recovery of product, centrifuge the original vials after thawing and prior to removing the cap.||Assay Summary:|1. Wash plate 2 times before adding standards and samples to wells!|2. Add 100ul standard or sample to each well and incubate for 90 minutes at 37°C. Aspirate. Do not wash plate!|3. Add 100ul Antibody (Biotin) working solution to each well and incubate for 60 minutes at 37°C. |4. Aspirate and wash 3 times.|5. Add 100ul SABC working solution to each well. Incubate for 30 minutes at 37°C|6. Aspirate and wash 5 times.|7. Add 90ul TMB substrate. Incubate 15-30 minutes at 37°C|8. Add 50ul Stop Solution. Read at 450nm immediately.|9. Calculate results.