Human Phosphoinositide-3-Kinase Class-2-Alpha Polypeptide (PIK3C2a) ELISA Kit

Principle of the Assay

The microtiter plate provided in this kit has been pre-coated with an antibody specific to PIK3C2a. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody preparation specific to PIK3C2a. 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 PIK3C2a, 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 PIK3C2a 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

Generates phosphatidylinositol 3-phosphate (PtdIns3P) and phosphatidylinositol 3,4-bisphosphate (PtdIns(3,4)P2) that act as second messengers. Has a role in several intracellular trafficking events. Functions in insulin signaling and secretion. Required for translocation of the glucose transporter SLC2A4/GLUT4 to the plasma membrane and glucose uptake in response to insulin-mediated RHOQ activation. Regulates insulin secretion through two different mechanisms: involved in glucose-induced insulin secretion downstream of insulin receptor in a pathway that involves AKT1 activation and TBC1D4/AS160 phosphorylation, and participates in the late step of insulin granule exocytosis probably in insulin granule fusion. Synthesizes PtdIns3P in response to insulin signaling. Functions in clathrin-coated endocytic vesicle formation and distribution. Regulates dynamin-independent endocytosis, probably by recruiting EEA1 to internalizing vesicles. In neurosecretory cells synthesizes PtdIns3P on large dense core vesicles. Participates in calcium induced contraction of vascular smooth muscle by regulating myosin light chain (MLC) phosphorylation through a mechanism involving Rho kinase-dependent phosphorylation of the MLCP-regulatory subunit MYPT1. May play a role in the EGF signaling cascade. May be involved in mitosis and UV-induced damage response. Required for maintenance of normal renal structure and function by supporting normal podocyte function. Involved in the regulation of ciliogenesis and trafficking of ciliary components (PubMed:31034465).

GENE ID	5286
SWISS PROT	O00443
SYNONYMS	PI3K2a; CPK; PI3-K-C2(Alpha); PI3-K-C2A; PI3K2-a; Phosphoinositide 3-Kinase 2a; Phosphatidylinositol 4-phosphate 3-kinase C2 domain-containing subunit alpha

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 PIK3C2a were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays): 3 samples with low, middle, and high-level PIK3C2a 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 1.6U/L.
ASSAY RANGE	3.12-200U/L
SPECIFICITY	This assay has high sensitivity and excellent specificity for the detection of PIK3C2a. No significant cross-reactivity or interference between PIK3C2a and analogs was observed. Note: Limited by current skills and knowledge, it is impossible to perform all possible cross-reactivity detection tests between PIK3C2a and all analogs, therefore, cross reactivity may still exist.