| Lumican, also known as LUM, is a protein which in humans is encoded by the LUM gene. This gene is a member of the small interstitial proteoglycan gene(SIPG) family, and it is a keratan sulfate proteoglycan which presents large quantities in the corneal stroma and in interstitial collagenous matrices of the heart, aorta, skeletal muscle, skin, and intervertebral discs. Lumican is mapped to 12q21.33, it interacts with collagen and limits growth of fibrils in diameter. In the cornea, Lumican not only interacts with collagen molecules to limit fibril growth, but also plays a critical role in the regular spacing of fibrils and acquisition of corneal transparency by virtue of its keratan sulfate-containing glycosaminoglycan side chains LDC. The Lumican, BioAssay(TM) ELISA Kit is for the quantitative detection of rat Lumican in cell culture supernates, serum and plasma (Heparin). Detection Range: 156-10,000pg/ml Sensitivity: <10pg/ml Specificity: Recognizes natural and recombinant rat Lumican. There is no detectable cross-reactivity with other relevant proteins. Test Principle: The Lumican, BioAssay(TM) ELISA Kit is a solid-phase immunoassay specially designed to measure rat Lumican with a 96-well strip plate that is pre-coated with antibody specific for Lumican. The detection antibody is a biotinylated antibody specific for Lumican. The capture antibody is monoclonal antibody from mouse and the detection antibody is polyclonal antibody from goat. The kit contains recombinant rat Lumican with immunogen: Expression system for standard: NS0 cells, Immunogen sequence: Q19-N338. To measure rat Lumican, add standards and samples to the wells, then add the biotinylated detection antibody. Wash the wells with PBS buffer, and add Avidin-Biotin-Peroxidase Complex (ABC-HRP). Wash away the unbounded ABC-HRP with PBS buffer and add TMB. TMB is substrate to HRP and will be catalyzed to produce a blue color product, which changes into yellow after adding acidic stop solution. The density of the yellow product is linearly proportional to rat Lumican in the sample. Read the density of the yellow product in each well using a plate reader, and benchmark the sample wells readings against the standard curve to determine the concentration of rat Lumican in the sample. Kit Components: 352965A: Microtiter Strips, 1x96 wells *352965B: Lumican Standard, 2x10ng 352965C: Anti-rat Lumican (Biotin), 1x100ul (1:100 dilution) 143696: Avidin-Biotin-Peroxidase Complex (ABC), 1x100ul (1:100 dilution) 143697: Sample Diluent Buffer, 1x30ml 143698: Antibody Diluent Buffer, 1x12ml 143699: ABC Diluent Buffer, 1x12ml 143700: TMB Color Developing Agent, 1x10ml 143701: TMB Stop Solution, 2N H2SO4, 1x10ml 143702: Wash Buffer, 25X, 1x20ml Storage and Stability: Store *352965B powder at 4C. Once reconstituted store at 4C for up to 12 hours or at -20C for up to 48 hours. Store other components at 4C. Stable for 6 months after receipt. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Assay Summary: 1. Add 100ul of samples and standards and incubate the plate at 37C for 90 minutes or at RT for 2 hours. Do not wash. 2. Add 100ul biotinylated antibody and incubate the plate at 37C for 60 minutes or at RT for 90 minutes. Wash plate 3 times with Working Wash Buffer. 3. Add 100ul of ABC working solution and incubate the plate at 37C for 30 minutes or at RT for 40 minutes. Wash plate 5 times with Working Wash Buffer. 4. Add 90ul of TMB color developing agent and incubate the plate at 37C in dark for 15-25 minutes or at RT for 30 minutes. 5. Add 100ul TMB Stop Solution and read. |
