Lysyl oxidase homolog 2 is an enzyme that in humans is encoded by the LOXL2 gene. This gene encodes a member of the lysyl oxidase gene family. The prototypic member of the family is essential to the biogenesis of connective tissue, encoding an extracellular copper-dependent amine oxidase that catalyses the first step in the formation of crosslinks in collagens and elastin. A highly conserved amino acid sequence at the C-terminus end appears to be sufficient for amine oxidase activity, suggesting that each family member may retain this function. The N-terminus is poorly conserved and may impart additional roles in developmental regulation, senescence, tumor suppression, cell growth control, and chemotaxis to each member of the family. LOXL2 can also crosslink collagen type IV and hence influence the sprouting of new blood vessels. Sample Type: Cell culture supernates, cell lysates, serum and plasma (heparin, EDTA). Intended Use: For quantitative detection of human LOXL2 in cell culture supernates, cell lysates, serum and plasma (heparin, EDTA). Sensitivity: <10pg/ml Range: 156-10,000pg/ml Specificity: Natural and recombinant human LOXL2 Test Principle: This human LOXL2 BioAssay(TM) ELISA Kit was based on standard sandwich enzyme-linked immune-sorbent assay technology. A monoclonal antibody from mouse specific for LOXL2 has been precoated onto 96-well plates. Standards (NSO,Q26-Q774) and test samples are added to the wells, a biotinylated detection polyclonal antibody from goat specific for LOXL2 is added subsequently and then followed by washing with PBS or TBS buffer. Avidin-Biotin-Peroxidase Complex was added and unbound conjugates were washed away with PBS or TBS buffer. HRP substrate TMB was used to visualize HRP enzymatic reaction. TMB was catalyzed by HRP to produce a blue color product that changed into yellow after adding acidic stop solution. The density of yellow is proportional to the human LOXL2 amount of sample captured in plate. Kit Components: 352962A: Microtiter Strips, 1x96 wells *352962B: LOXL2 Standard, 2x1 vial Recombinant human LOXL2 352962C: anti human LOXL2 (Biotin), 1x130ul 143696: Avidin-Biotin-Peroxidase Complex (ABC), 1x130ul (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 Storage and Stability: Store *352962B 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. Aspirate the liquid from wells. 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 0.01M PBS or TBS. 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 0.01M PBS or TBS. 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.
