Rat Hypoxia Inducible Factor 1 Alpha (HIF1a) ELISA Kit

Principle of the Assay

The microtiter plate provided in this kit has been pre-coated with an antibody specific to HIF1a. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated antibody preparation specific to HIF1a. 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 HIF1a, 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 HIF1a 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

Functions as a master transcriptional regulator of the adaptive response to hypoxia. Under hypoxic conditions, activates the transcription of over 40 genes, including erythropoietin, glucose transporters, glycolytic enzymes, vascular endothelial growth factor, HILPDA, and other genes whose protein products increase oxygen delivery or facilitate metabolic adaptation to hypoxia. Plays an essential role in embryonic vascularization, tumor angiogenesis and pathophysiology of ischemic disease (By similarity). Heterodimerizes with ARNT; heterodimer binds to core DNA sequence 5'-TACGTG-3' within the hypoxia response element (HRE) of target gene promoters (By similarity). Activation requires recruitment of transcriptional coactivators such as CREBBP and EP300. Activity is enhanced by interaction with NCOA1 and/or NCOA2. Interaction with redox regulatory protein APEX1 seems to activate CTAD and potentiates activation by NCOA1 and CREBBP. Involved in the axonal distribution and transport of mitochondria in neurons during hypoxia (By similarity).

GENE ID	29560
SWISS PROT	O35800
SYNONYMS	HIF1-A; MOP1; PASD8; Basic Helix-Loop-Helix Transcription Factor; ARNT-interacting protein; Basic-helix-loop-helix-PAS protein MOP1; Class E basic helix-loop-helix protein 78

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 HIF1a were tested 20 times on one plate, respectively. Inter-assay Precision (Precision between assays): 3 samples with low, middle, and high-level HIF1a 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 0.057ng/mL.
ASSAY RANGE	0.156-10ng/mL
SPECIFICITY	This assay has high sensitivity and excellent specificity for the detection of HIF1a. No significant cross-reactivity or interference between HIF1a and analogs was observed. Note: Limited by current skills and knowledge, it is impossible to perform all possible cross-reactivity detection tests between HIF1a and all analogs, therefore, cross reactivity may still exist.