Rat RalA Binding Protein 1 (RALBP1) ELISA Kit

Principle of the Assay

The microtiter plate provided in this kit has been pre-coated with an antibody specific to RALBP1. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody preparation specific to RALBP1. 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 RALBP1, 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 RALBP1 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

Multifunctional protein that functions as a downstream effector of RALA and RALB. As a GTPase-activating protein/GAP can inactivate CDC42 and RAC1 by stimulating their GTPase activity (PubMed:7623849, PubMed:9422736). As part of the Ral signaling pathway, may also regulate ligand-dependent EGF and insulin receptors-mediated endocytosis (PubMed:10393179). During mitosis, may act as a scaffold protein in the phosphorylation of EPSIN/EPN1 by the mitotic kinase cyclin B-CDK1, preventing endocytosis during that phase of the cell cycle. During mitosis, also controls mitochondrial fission as an effector of RALA. Recruited to mitochondrion by RALA, acts as a scaffold to foster the mitotic kinase cyclin B-CDK1-mediated phosphorylation and activation of DNM1L (By similarity).

GENE ID	84014
SWISS PROT	Q62796
SYNONYMS	RIP; RIP1; RLIP76; 76 kDa Ral-interacting protein; Ral-interacting protein 1; Dinitrophenyl S-glutathione ATPase

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 RALBP1 were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays): 3 samples with low, middle, and high-level RALBP1 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 0.058ng/mL.
ASSAY RANGE	0.156-10ng/mL
SPECIFICITY	This assay has high sensitivity and excellent specificity for the detection of RALBP1. No significant cross-reactivity or interference between RALBP1 and analogs was observed. Note: Limited by current skills and knowledge, it is impossible to perform all possible cross-reactivity detection tests between RALBP1 and all analogs, therefore, cross reactivity may still exist.