Sample Type: Serum, plasma, tissue homogenates Intended Use: For the quantitative determination of human lipoprotein-associated phospholipase A2 (Lp-PLA2) concentrations in serum, plasma, tissue homogenates. Sensitivity: 0.39ng/ml Range: 1.56-100ng/ml Specificity: This assay has high sensitivity and excellent specificity for detection of human Lp-PLA2. No significant cross-reactivity or interference between human Lp-PLA2 and analogues was observed. Principle of the Assay: This assay employs the quantitative sandwich enzyme immunoassay technique. Antibody specific for Lp-PLA2 has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and any Lp-PLA2 present is bound by the immobilized antibody. After removing any unbound substances, a biotin-conjugated antibody specific for Lp-PLA2 is added to the wells. After washing, avidin conjugated Horseradish Peroxidase (HRP) is added to the wells. Following a wash to remove any unbound avidin-enzyme reagent, a substrate solution is added to the wells and color develops in proportion to the amount of Lp-PLA2 bound in the initial step. The color development is stopped and the intensity of the color is measured. Kit Components: 351661A: Microtiter Strips, 12x8 Microwell Strips. Pre-coated, ready to use. 351661B: Standard (lyophilized), 2 vials 351661C: Biotin-Antibody (100x Concentrate), 1x120ul 351661D: HRP-Avidin, (100x Concentrate), 1x120ul 351661E: Biotin-Antibody Diluent, 1x15ml 351661F: HRP-Avidin Diluent, 1x15ml 351661G: Sample Diluent, 1x50ml 351661H: Wash Buffer (25X Concentrate), 1x20ml 351661J: TMB Substrate, 1x10ml 351661K: Stop Solution, 1x10ml Storage and Stability: Store all kit components at 4C. Unopened kit is stable for 6 months after receipt. Once opened, the kit is stable for 1 month provided this is within the expiration date of the kit. Opened Kit: Coated plate may be stored at 4C for up to 1 month, sealed in the aluminum foil bag. Standard, Biotin-Antibody and HRP-Avidin may be stored at 4C or -20C for up to 1 month. The rest of the components may be stored at 4C for up to 1 month. For maximum recovery of contents, centrifuge the original vials after thawing and prior to removing the cap. Assay Summary: 1. Prepare reagents, standards and samples as instructed. 2. Add 100ul standard or sample to each well. Incubate for 2 hours at 37C 3. Remove the liquid of each wee, do not wash. 4. Add 100ul Biotin-Antibody (1x) to each well. Incubate for 1 hour at 37C 5. Aspirate and wash 3 times. 6. Add 100ul HRP-Avidin (1x) to each well. Incubate for 1 hour at 37C 7. Aspirate and wash 5 times. 8. Add 90ul TMB substrate to each well Incubate for 15-30 minutes at 37C. Protect from light. 9. Add 50ul Stop Solution to each well. Read at 450nm within 5 minutes.
