Porcine S100 Calcium Binding Protein A12 (S100A12) ELISA Kit

Principle of the Assay

The microtiter plate provided in this kit has been pre-coated with an antibody specific to S100A12. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody preparation specific to S100A12. Next, Avidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After the TMB substrate solution is added, only those wells that contain S100A12, biotin-conjugated antibody, and enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of sulphuric acid solution, and the color change is measured spectrophotometrically at a wavelength of 450nm ± 10nm. The concentration of S100A12 in the samples is then determined by comparing the O.D. of the samples to the standard curve.

For Use with serum, plasma, and cell culture supernatants. For Research Use Only. Not for use in diagnostic procedures.

Target Information

S100A12 is a calcium-, zinc- and copper-binding protein which plays a prominent role in the regulation of inflammatory processes and immune response. Its pro-inflammatory activity involves recruitment of leukocytes, promotion of cytokine and chemokine production, and regulation of leukocyte adhesion and migration. Acts as an alarmin or a danger associated molecular pattern (DAMP) molecule and stimulates innate immune cells via binding to receptor for advanced glycation endproducts (AGER). Binding to AGER activates the MAP-kinase and NF-kappa-B signaling pathways leading to production of pro-inflammatory cytokines and up-regulation of cell adhesion molecules ICAM1 and VCAM1. Acts as a monocyte and mast cell chemoattractant. Can stimulate mast cell degranulation and activation which generates chemokines, histamine and cytokines inducing further leukocyte recruitment to the sites of inflammation. Can inhibit the activity of matrix metalloproteinases; MMP2, MMP3 and MMP9 by chelating Zn(2+) from their active sites (By similarity).

GENE ID	100301483
SWISS PROT	P80310
SYNONYMS	CGRP; P6; CAAF1; CAGC; ENRAGE; MRP6; Calgranulin C; Calcium-binding protein in amniotic fluid 1; Extracellular newly identified RAGE-binding; Migration inhibitory factor-related 6

Materials Supplied

Kit Components	96 Wells Quantity/Size
Pre-coated, ready-to-use 96-well strip plate	1 plate
Plate sealer for 96 wells	2
Standard	2 tubes
Diluent buffer	1 bottle
Detection Reagent A	1 bottle
Detection Reagent B	1 bottle
TMB Substrate	1 tube
Stop Solution	1 tube
Wash Buffer (30 ℅ concentrate)	1 tube
Product data sheet	1 copy

Storage

The TMB Substrate, Wash Buffer (30X concentrate), and the Stop Solution should be stored at 4°C upon receipt, while the other items should be stored at -20°C.

Performance Characteristics

REPEATABILITY	Intra-assay Precision (Precision within an assay): 3 samples with low, middle, and high-level S100A12 were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays): 3 samples with low, middle, and high-level S100A12 were tested on 3 different plates, with 8 replicates in each plate. CV(%) = SD/meanX100 Intra-Assay: CV<10% Inter-Assay: CV<12%
SENSITIVITY	The minimum detectable dose was 0.043ng/mL.
ASSAY RANGE	0.156-10ng/mL
SPECIFICITY	This assay has high sensitivity and excellent specificity for the detection of S100A12. No significant cross-reactivity or interference between S100A12 and analogs was observed. Note: Limited by current skills and knowledge, it is impossible to perform all possible cross-reactivity detection tests between S100A12 and all analogs, therefore, cross reactivity may still exist.