Mouse Insulin Degrading Enzyme (IDE) ELISA Kit

Principle of the Assay

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

Plays a role in the cellular breakdown of insulin, APP peptides, IAPP peptides, natriuretic peptides, glucagon, bradykinin, kallidin, and other peptides, and thereby plays a role in intercellular peptide signaling (PubMed:9830016, PubMed:12634421, PubMed:12732730, PubMed:24847884, PubMed:26394692). Substrate binding induces important conformation changes, making it possible to bind and degrade larger substrates, such as insulin (By similarity). Contributes to the regulation of peptide hormone signaling cascades and regulation of blood glucose homeostasis via its role in the degradation of insulin, glucagon and IAPP (PubMed:24847884, PubMed:26394692). Plays a role in the degradation and clearance of APP-derived amyloidogenic peptides that are secreted by neurons and microglia (PubMed:9830016). Degrades the natriuretic peptides ANP, BNP and CNP, inactivating their ability to raise intracellular cGMP (By similarity). Also degrades an aberrant frameshifted 40-residue form of NPPA (fsNPPA) which is associated with familial atrial fibrillation in heterozygous patients (By similarity). Involved in antigen processing. Produces both the N terminus and the C terminus of MAGEA3-derived antigenic peptide (EVDPIGHLY) that is presented to cytotoxic T lymphocytes by MHC class I.

GENE ID	15925
SWISS PROT	Q9JHR7
SYNONYMS	Insulysin; Insulin Protease; Abeta-degrading protease; Insulinase

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 IDE were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays): 3 samples with low, middle, and high-level IDE 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 1.33pg/mL.
ASSAY RANGE	3.12-200pg/mL
SPECIFICITY	This assay has high sensitivity and excellent specificity for the detection of IDE. No significant cross-reactivity or interference between IDE and analogs was observed. Note: Limited by current skills and knowledge, it is impossible to perform all possible cross-reactivity detection tests between IDE and all analogs, therefore, cross reactivity may still exist.