Human HtrA Serine Peptidase 1 (HTRA1) ELISA Kit

Principle of the Assay

The microtiter plate provided in this kit has been pre-coated with an antibody specific to HTRA1. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody preparation specific to HTRA1. 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 HTRA1, 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 HTRA1 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

Serine protease with a variety of targets, including extracellular matrix proteins such as fibronectin. HTRA1-generated fibronectin fragments further induce synovial cells to up-regulate MMP1 and MMP3 production. May also degrade proteoglycans, such as aggrecan, decorin and fibromodulin. Through cleavage of proteoglycans, may release soluble FGF-glycosaminoglycan complexes that promote the range and intensity of FGF signals in the extracellular space. Regulates the availability of insulin-like growth factors (IGFs) by cleaving IGF-binding proteins. Inhibits signaling mediated by TGF-beta family members. This activity requires the integrity of the catalytic site, although it is unclear whether TGF-beta proteins are themselves degraded. By acting on TGF-beta signaling, may regulate many physiological processes, including retinal angiogenesis and neuronal survival and maturation during development. Intracellularly, degrades TSC2, leading to the activation of TSC2 downstream targets.

GENE ID	5654
SWISS PROT	Q92743
SYNONYMS	HtrA; L56; ORF480; PRSS11; HtrA Serine Peptidase 1; Protease,Serine,11; IGFBP5-Protease; High-temperature requirement A serine peptidase 1

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 HTRA1 were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays): 3 samples with low, middle, and high-level HTRA1 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 13.2pg/mL.
ASSAY RANGE	31.2-2000pg/mL
SPECIFICITY	This assay has high sensitivity and excellent specificity for the detection of HTRA1. No significant cross-reactivity or interference between HTRA1 and analogs was observed. Note: Limited by current skills and knowledge, it is impossible to perform all possible cross-reactivity detection tests between HTRA1 and all analogs, therefore, cross reactivity may still exist.