Rat hypoxia-inducible factor 1α,HIF-1α ELISA Kit

Instructions
Code CSB-E08540r
Size 96T,5×96T,10×96T
Trial Size 24T ELISA kits trial application
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Product Details

Description

The Rat hypoxia-inducible factor 1α (HIF-1α) ELISA kit is a ready-to-use microwell, strip plate ELISA kit for the quantitative measurement of rat HIF-1α in biological samples, including serum, plasma, and tissue homogenates. It employs the sandwich ELISA technique and enzyme-substrate chromogenic reaction in the detection of target analytes in the samples. This ELISA kit has been quality controlled in multiple characteristics, including sensitivity, specificity, precision, linearity, and recovery. See the product instructions to obtain more information.

HIF-1α, a basic-helix-loop-helix (bHLH)-PER (Period)-ARNT-SIM (single-minded) (PAS) superfamily member, is the O2-labile HIF-1 subunit and is widely expressed in hypoxia. HIF-1α orchestrates cellular adaptation to low oxygen and nutrient-deprived environment and drives progression to malignancy in human solid cancers. Under normal O2 tension, the prolyl hydroxylases (PHDs) become active and hydroxylate HIF-1α, triggering von Hippel–Lindau (pVHL)-mediated ubiquitination and proteasomal degradation HIF-1. Hypoxia inhibits the enzymatic activity of PHDs, terminating proline modification and pVHL-HIF binding, causing HIF-1α stabilization, nuclear translocation, and further formation of HIF-1 complex. This cascade of events ultimately induces the transcription of genes that promote glycolytic metabolism, angiogenesis, and survival. HIF-1α is physiologically activated during embryogenesis and in wound-healing processes. Whereas in cancer, HIF-1α activation is associated with malignancy and poor prognosis.

Target Name hypoxia inducible factor 1, alpha subunit (basic helix-loop-helix transcription factor)
Alternative Names Hif1aHypoxia-inducible factor 1-alpha ELISA Kit; HIF-1-alpha ELISA Kit; HIF1-alpha ELISA Kit
Abbreviation HIF1A
Uniprot No. O35800
Species Rattus norvegicus (Rat)
Sample Types serum, plasma, tissue homogenates
Detection Range 3.12 pg/mL-200 pg/mL
Sensitivity 0.78 pg/mL
Assay Time 1-5h
Sample Volume 50-100ul
Detection Wavelength 450 nm
Research Area Epigenetics and Nuclear Signaling
Assay Principle quantitative
Measurement Sandwich
Precision
Intra-assay Precision (Precision within an assay): CV%<8%
Three samples of known concentration were tested twenty times on one plate to assess.
Inter-assay Precision (Precision between assays): CV%<10%
Three samples of known concentration were tested in twenty assays to assess.
Linearity
To assess the linearity of the assay, samples were spiked with high concentrations of rat HIF-1α in various matrices and diluted with the Sample Diluent to produce samples with values within the dynamic range of the assay.
 SampleSerum(n=4)
1:1Average %90
Range %87-94
1:2Average %96
Range %91-100
1:4Average %93
Range %87-97
1:8Average %100
Range %96-103
Recovery
The recovery of rat HIF-1α spiked to levels throughout the range of the assay in various matrices was evaluated. Samples were diluted prior to assay as directed in the Sample Preparation section.
Sample TypeAverage % RecoveryRange
Serum (n=5) 9287-95
EDTA plasma (n=4)9588-98
Typical Data
These standard curves are provided for demonstration only. A standard curve should be generated for each set of samples assayed.
pg/mlOD1OD2AverageCorrected
2002.715 2.656 2.686 2.538
1001.730 1.676 1.703 1.555
501.196 1.179 1.188 1.040
250.608 0.618 0.613 0.465
12.50.404 0.412 0.408 0.260
6.250.303 0.316 0.310 0.162
3.120.248 0.237 0.243 0.095
00.150 0.146 0.148  
Materials provided
  • A micro ELISA plate ---The 96-well plate has been pre-coated with an anti-Rat HIF-1α antibody. This dismountable microplate can be divided into 12 x 8 strip plates.
  • Two vials lyophilized standard ---Dilute a bottle of the standard at dilution series, read the OD values, and then draw a standard curve.
  • One vial Biotin-labeled HIF-1α antibody (100 x concentrate) (120 μl/bottle) ---Act as the detection antibody.
  • One vial HRP-avidin (100 x concentrate) (120 μl/bottle) ---Bind to the detection antibody and react with the TMB substrate to make the solution chromogenic.
  • One vial Biotin-antibody Diluent (15 ml/bottle) ---Dilute the Biotin-antibody.
  • One vial HRP-avidin Diluent (15 ml/bottle) ---Dilute the HRP-avidin solution.
  • One vial Sample Diluent (50 ml/bottle)---Dilute the sample to an appropriate concentration.
  • One vial Wash Buffer (25 x concentrate) (20 ml/bottle) ---Wash away unbound or free substances.
  • One vial TMB Substrate (10 ml/bottle) ---Act as the chromogenic agent. TMB interacts with HRP, eliciting the solution turns blue.
  • One vial Stop Solution (10 ml/bottle) ---Stop the color reaction. The solution color immediately turns from blue to yellow.
  • Four Adhesive Strips (For 96 wells) --- Cover the microplate when incubation.
  • An instruction manual
Materials not provided
  • A microplate reader capable of measuring absorbance at 450 nm, with the correction wavelength set at 540 nm or 570 nm.
  • An incubator can provide stable incubation conditions up to 37°C±5°C.
  • Centrifuge
  • Vortex
  • Squirt bottle, manifold dispenser, or automated microplate washer
  • Absorbent paper for blotting the microtiter plate
  • 50-300ul multi-channel micropipette
  • Pipette tips
  • Single-channel micropipette with different ranges
  • 100ml and 500ml graduated cylinders
  • Deionized or distilled water
  • Timer
  • Test tubes for dilution
Troubleshooting
and FAQs
ELISA kit FAQs
Storage Store at 2-8°C. Please refer to protocol.
Lead Time 3-5 working days

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Target Background

Function
(From Uniprot)
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. Binds to core DNA sequence 5'-[AG]CGTG-3' within the hypoxia response element (HRE) of target gene promoters. Activation requires recruitment of transcriptional coactivators such as CREBBP and EP300. Activity is enhanced by interaction with both, NCOA1 or NCOA2. Interaction with redox regulatory protein APEX 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 References into Functions
  1. Serum level of cardiac biomarkers and the necrotic areas were significantly lower in the Ad-HIF-1a-Trip group compared to the other groups in myocardial infarction. Cardiac function in the Ad-HIF-1alpha-Trip group was decreased less profoundly through day 7 to day 28 compared to the other groups. PMID: 30423574
  2. the interplay between HIF1alpha and ROCK1 may be a critical factor that regulates cell proliferation and collagen synthesis in rat hepatic stellate cells under hypoxia. PMID: 30132575
  3. Early changes in impedance in response to I/R is related to histopathological changes, the nuclear stabilization and translocation of HIF-1alpha as well as expression of iNOS. PMID: 29451295
  4. Stud shows that HIF-1alpha induction during reperfusion is a protector mechanism implicated in a normal renal tissue repair after ischemia/reperfusion. PMID: 28106131
  5. HRS reduced mechanical hyperalgesia and activation of cell autophagy in neuropathic pain through a HIF1-dependent pathway. PMID: 29888265
  6. Succinate acts as a signaling molecule in hypoxic stabilization and HIF-1alpha expression induction in the brain cortex. PMID: 29308570
  7. pathologic hypoxia stimulates neural stem cells proliferation by increasing expression of HIF-1alpha and activating the Wnt/beta-catenin signaling pathway. PMID: 28838333
  8. These results indicate that HIF-1alpha expression can be induced and activated in rats during the acute lung inflammatory damages triggered by septic lymph injection and that lung inflammatory injuries occur via a HIF-1alpha-dependent pathway. PMID: 28478272
  9. EC-mediated protection is only possible when there is a bidirectional crosstalk between ECs and CMs even without physical cell-cell contact. In addition, this protective effect is at least partially related to cell-ECM interactions and HIF-1alpha, which is regulated by HIF1A-AS1 under oxidative stress. PMID: 28629892
  10. Data suggest that overexpression of hypoxia inducible factor 1 alpha subunit (HIF-1alpha) by gene therapy may be a useful method for enhancing alveolar bone defect osteogenesis and angiogenesis. PMID: 26929250
  11. By using a cell model of desferoxamine-dependent HIF activation, we demonstrated that propofol was able to inhibit apoptosis and mitochondrial depolarization associated to HIF-1a action. In conclusion, hepatic I/R injury induces mitochondrial dysfunction that is not prevented by inhaled sevoflurane. PMID: 27154980
  12. Hypoxia and hyperoxia differentially control proliferation of rat neural crest stem cells via distinct regulatory pathways of the HIF1alpha-CXCR4 and TP53-TPM1 proteins PMID: 28002632
  13. HDACs controlled HIF-1alpha stability by regulating HIF-1alpha-PHD2 interaction in NP cells. PMID: 26765925
  14. The GO-Cu nanocomposites upregulate the expression of Hif-1alpha in BMSCs. PMID: 26945787
  15. Impaired ischemia-induced angiogenesis in rats with chronic kidney disease can be improved by HIF stabilization, even if started after onset of ischemia. PMID: 27927598
  16. Warburg effect plays a critical role in PDGF-induced pulmonary arterial smooth muscle cells proliferation and is mediated by activation of the PI3K signaling pathway and HIF-1alpha. PMID: 28738389
  17. Work identifies HIF-1 as an early chronic kidney disease (CKD)-related pathological event, prospective marker, and potential target against vascular calcification in CKD-relevant conditions. PMID: 27470678
  18. High HIF1A expression is associated with inflammation and fibrosis in kidney. PMID: 26891083
  19. Hypoxia-inducible factor 1alpha knockdown could be a potential approach to improve the efficacy of temozolomide therapy for pituitary adenomas. PMID: 26647409
  20. Report significant impairment of renal oxygenation and mitochondrial function at the early stages of chronic kidney disease and demonstrate the beneficial role of HIF-1alpha activation on renal function and metabolism. PMID: 28331062
  21. We show that HACT requires transcriptional activation of HIF-1alpha throughout the course of HA and that this activation is accompanied by two metabolic switches. PMID: 28274939
  22. In response to hypoxia, compared with rats, mice had higher minute ventilation, lower metabolic rate and higher expression of HIF-1alpha in the brainstem. PMID: 27742895
  23. Can use light microscopy to separate renal cells into those with and without HIF1alpha expression. PMID: 28212880
  24. HIF-1alpha activation via cytosolic Ca(2+) signaling pathway play a role in the mechanism of isoflurane-induced neurodegeneration in neonatal rodents, suggesting HIF-1alpha as a potential therapeutic target for the prevention of the deleterious effects of prolonged exposures to anesthetics. PMID: 27769790
  25. Immunohistochemistry staining, mRNA, and protein for hypoxia-inducible factor 1alpha were upregulated within the paraventricular nucleus of left coronary artery-ligated chronic heart failure rats. PMID: 27810863
  26. The expression of three well-defined downstream targets of hypoxia inducible factor 1 alpha subunit (HIF-1alpha), pyruvate dehydrogenase kinase 1 (PDK1), lactate dehydrogenase A (LDHA) and vascular endothelial growth factor (VEGF), were up-regulated in hippocampus after 28 days of simulated microgravity exposure. PMID: 27988334
  27. The mechanism of development of liver fibrosis involves activation of matrix metalloproteinase-2 and matrix metalloproteinase-9, up-regulation of HIF-1alpha transcriptional activity and its related factors, NF-kappaB and TGF-beta. PMID: 27260943
  28. Reduction of endoplasmic reticulum stress by 4-phenylbutyric acid reduces chronic hypoxia induced right ventricular hypertrophy and improves hypoxic tolerance through upregulation of HIF-1alpha. PMID: 27058435
  29. Nox2 is a modulator of CsA-induced hypoxia upstream of HIF-1alpha PMID: 26950727
  30. The results demonstrate that inflammation-induced fetal growth restriction is associated with increased placental HIF-1alpha accumulation; however, expression of this transcription factor may not correlate with regions of hypoxia in late-gestation placentas. PMID: 28423052
  31. this study demonstrates the potent suppressive effects of HIF-1alpha shRNA on hypoxia-induced PH and PASMC hyperplasia, providing evidence for the potential application of HIF-1alpha shRNA in the treatment of hypoxic PH. PMID: 27748831
  32. The present experiments indicated that IQ enhanced the radiosensitivity of C6 rat glioma cells both in vitro and in vivo. The primary mechanism of this radiosensitizing effect involves the downregulation of HIF-1alpha. F-FDG and F-FMISO PET/CT were sensitive and noninvasive for monitoring the early radiosensitizing effect of IQ. Meanwhile, F-FMISO PET/CT provided more information on the changes in tumor hypoxic status. PMID: 26963468
  33. Stabilization of HIF-1alpha by inhibiting HIF-1alpha hydroxylation prevented the ischemic decrease in both PMCA1 and PMCA2 mRNAs in CA1 neurons PMID: 26969192
  34. Data show that cardiac arrest (CA) increased HIF-1alpha and pro-inflammatory cytokines (PICs) including IL-6 and TNF-alpha in the hippocampus CA1 region. PMID: 27383646
  35. specific signaling pathways engaged in the development of diabetic retinopathy, including the activation of mTOR and HIF-1alpha -VEGF mechanism, were investigated. PMID: 27997905
  36. Hypoxia could cause HIF-1alpha-dependent increase in the expression of p27 leading to cell cycle arrest in G0/G1 phase PMID: 26920732
  37. hypoxia or GSN overexpression induces HIF-1alpha expression and reduces the expression of survival markers p-Akt and Bcl-2 in H9c2 cardiomyoblast cells PMID: 27193608
  38. HIF-1A regulates epithelial-mesenchymal transition via the Snail and beta-catenin pathways in paraquat poisoning-induced early pulmonary fibrosis. PMID: 26781174
  39. Data show that under hypoxia, sirtuin 1 (SIRT1) expression was raised and matrix metalloproteinase-9 (MMP-9) and hypoxia inducible factor 1alpha (HIF-1alpha) were reduced by Lycium barbarum polysaccharides (LBP) in a dose-dependent manner. PMID: 27363270
  40. tumor necrosis factor tumor necrosis factor cachectin tumor necrosis factor (TNF superfamily member 2) tumor necrosis factor alpha tumor necrosis factor ligand superfamily member 2 PMID: 26614261
  41. Chronic intermittent hypobaric hypoxia has renal protective effects in diabetic rats, which might be related with activating HIF1alpha signaling. PMID: 27351799
  42. shRNA of HIF-1alpha could antagonize the hypoxia-mediated increase in hepatic cancer cell migration and invasion, and synergize with hypoxia to inhibit the cell proliferation in HCC cells PMID: 25388848
  43. reduced miR-195 expression partially protects rats from spinal cord injury, primarily by targeting HIF-1alpha. PMID: 26927342
  44. 24 h after hypoxic-ischemic injury, there was a significant increase in PHD3 protein and an increase of HIF-1alpha compared to controls PMID: 26108712
  45. nano-microcapsule bFGF combined with HIF-1 prevented random skin flap survival in rats through antioxidative, anti-inflammatory and activation of the protein expression levels of COX-2 and VEGF. PMID: 26707180
  46. Study suggested that neonatal bronchopulmonary dysplasia caused impaired cognitive function and neuron apoptosis in hippocampus via p53 and HIF-1alpha PMID: 26431790
  47. Coupled with HIF-1alpha and vascular endothelial growth factor (VEGF). PMID: 26537366
  48. expression of VEGF and HIF-1alpha might be involved with myocardial remodeling and angiogenesis in myocardial infarction PMID: 26813461
  49. HIF1alphamediated expression of VEGF may be one of the important mechanisms regulating ovarian luteal development in mammals in vivo. PMID: 26323652
  50. The hypoxia-induced HIF-1alpha could promote glucose metabolism to protect hepatocellular mitochondria from damage. It could be a useful way to protect liver against I/R injuries and inflammatory injury. PMID: 25726501

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Subcellular Location Cytoplasm, Nucleus, Nucleus speckle
Tissue Specificity Expressed in the kidney, higher expression is seen in the renal medulla than in the cortex. Expressed also in the perivenous zone of the liver.
Database Links

KEGG: rno:29560

STRING: 10116.ENSRNOP00000042230

UniGene: Rn.10852

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