Human RAR alpha (Retinoic Acid Receptor Alpha) ELISA Kit
This ELISA kit uses Sandwich-ELISA as the method. The micro ELISA plate provided in this kit has been pre-coated with an antibody specific to Human RAR alpha . Standards or samples are added to the appropriate micro ELISA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Human RAR alpha and Avidin-Horseradish Peroxidase (HRP) conjugate are added to each micro plate well successively and incubated. Free components are washed away. The substrate solution is added to each well. Only those wells that contain Human RAR alpha, biotinylated detection antibody and Avidin-HRP conjugate will appear blue in color. The enzyme-substrate reaction is terminated by adding Stop Solution and the color turns yellow. The optical density (OD) is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The OD value is proportional to the concentration of Human RAR alpha. The concentration of Human RAR alpha in samples can be calculated by comparing the OD of the samples to the standard curve.
|Detection Range||0.16-10 ng/mL|
|Sample Volume Required Per Well||100uL|
|Sample Type||Serum, plasma and other biological fluids|
This kit recognizes Human RAR alpha in samples. No significant cross-reactivity or interference between Human RAR alpha and analogues was observed.
As the OD values of the standard curve may vary according to the conditions of the actual assay performance (e.g. operator, pipetting technique, washing technique or temperature effects), the operator should establish a standard curve for each test. Typical standard curve and data is provided below for reference only.
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human RAR alpha were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human RAR alpha were tested on 3 different plates, 20 replicates in each plate.
|Intra-assay Precision||Inter-assay Precision|
|C V (%)||5.66||5.00||3.89||7.02||4.96||4.16|
The recovery of Human RAR alpha spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.
|Sample Type||Range (%)||Average Recovery (%)|
|EDTA plasma (n=5)||87-99||94|
|Cell culture media (n=5)||96-107||101|
Samples were spiked with high concentrations of Human RAR alpha and diluted with Reference Standard & Sample Diluent to produce samples with values within the range of the assay.
|Serum (n=5)||EDTA plasma (n=5)||Cell culture media (n=5)|
Kit Components & Storage
An unopened kit can be stored at 4'C for 1 month. If the kit is not used within 1 month, store the items separately according to the following conditions once the kit is received.
|Micro ELISA Plate(Dismountable)||8 wells X12 strips||-20'C, 6 months|
|Reference Standard||2 vials|
|Concentrated Biotinylated Detection Ab (100X)||1 vial, 120 uL|
|Concentrated HRP Conjugate (100X)||1 vial, 120 uL||-20'C(shading light), 6 months|
|Reference Standard & Sample Diluent||1 vial, 20 mL||4'C, 6 months|
|Biotinylated Detection Ab Diluent||1 vial, 14 mL|
|HRP Conjugate Diluent||1 vial, 14 mL|
|Concentrated Wash Buffer (25X)||1 vial, 30 mL|
|Substrate Reagent||1 vial, 10 mL||4'C(shading light)|
|Stop Solution||1 vial, 10 mL||4'C|
|Plate Sealer||5 pieces|
|Product Description||1 copy|
|Certificate of Analysis||1 copy|
Human RAR alpha (Retinoic Acid Receptor Alpha) ELISA Kit (HUES03280) Assay procedure
- 1. Set standard, test sample and control (zero) wells on the pre-coated plate and record theirpositions. It is recommended to measure each standard and sample in duplicate. Note: addall solutions to the bottom of the plate wells while avoiding contact with the well walls. Ensuresolutions do not foam when adding to the wells.
- 2. Aliquot 100µl of standard solutions into the standard wells.
- 3. Add 100µl of Sample / Standard dilution buffer into the control (zero) well.
- 4. Add 100µl of properly diluted sample (serum, plasma, tissue homogenates and otherbiological fluids) into test sample wells.
- 5. Cover the plate with the sealer provided in the kit and incubate for 90 min at 37°C.
- 6. Aspirate the liquid from each well, do not wash. Immediately add 100µL of BiotinylatedDetection Ab working solution to each well. Cover the plate with a plate seal and gently mix.Incubate for 1 hour at 37°C.
- 7. Aspirate or decant the solution from the plate and add 350µL of wash buffer to each welland incubate for 1-2 minutes at room temperature. Aspirate the solution from each well andclap the plate on absorbent filter paper to dry. Repeat this process 3 times. Note: a microplatewasher can be used in this step and other wash steps.
- 8. Add 100µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37°C.
- 9. Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
- 10. Add 90µL of Substrate Reagent to each well. Cover with a new plate seal and incubate forapproximately 15 min at 37°C. Protect the plate from light. Note: the reaction time can beshortened or extended according to the actual color change, but not by more than 30min.
- 11. Add 50 µL of Stop Solution to each well. Note: Adding the stop solution should be done inthe same order as the substrate solution.
- 12. Determine the optical density (OD value) of each well immediately with a microplate readerset at 450 nm.
Human RAR alpha (Retinoic Acid Receptor Alpha) ELISA Kit (HUES03280) Protein Information
|UniProt Protein Function:||RARA: is a receptor for retinoic acid, a potent mammalian morphogen and teratogen that has profound effects on vertebrate development. RARA is a member of the nuclear receptor superfamily. Controls cell function by directly regulating gene expression. Its phosphorylation is crucial for transcriptional activity. Aberrations involving RARA may be a cause of acute promyelocytic leukemia. Two splice-variant isoforms have been described.|
|UniProt Protein Details:|
Protein type:DNA-binding; Oncoprotein; Transcription factor; Nuclear receptor
Chromosomal Location of Human Ortholog: 17q21
Cellular Component: nucleoplasm; cell surface; cell soma; perinuclear region of cytoplasm; cytoplasm; dendrite; nuclear chromatin; nucleus; actin cytoskeleton
Molecular Function:protein domain specific binding; protein kinase B binding; retinoic acid binding; zinc ion binding; chromatin DNA binding; translation repressor activity, nucleic acid binding; transcription coactivator activity; phosphoinositide 3-kinase regulator activity; drug binding; alpha-actinin binding; transcription factor binding; protein binding; enzyme binding; protein heterodimerization activity; protein kinase A binding; steroid hormone receptor activity; mRNA 5'-UTR binding; retinoic acid receptor activity; transcription factor activity; transcription corepressor activity; receptor binding
Biological Process: retinoic acid receptor signaling pathway; limb development; prostate gland development; negative regulation of translational initiation; regulation of myelination; estrogen receptor signaling pathway; glandular epithelial cell development; positive regulation of transcription, DNA-dependent; ventricular cardiac muscle cell differentiation; regulation of synaptic plasticity; female pregnancy; negative regulation of transcription from RNA polymerase II promoter; signal transduction; protein amino acid phosphorylation; regulation of phosphoinositide 3-kinase activity; response to estradiol stimulus; response to vitamin A; negative regulation of granulocyte differentiation; germ cell development; positive regulation of interleukin-4 production; negative regulation of cell proliferation; Sertoli cell fate commitment; positive regulation of T-helper 2 cell differentiation; ureteric bud development; negative regulation of interferon-gamma production; positive regulation of cell proliferation; positive regulation of interleukin-13 production; transmembrane transport; positive regulation of interleukin-5 production; transcription initiation from RNA polymerase II promoter; response to retinoic acid; multicellular organism growth; positive regulation of cell cycle; positive regulation of binding; negative regulation of tumor necrosis factor production; liver development; embryonic camera-type eye development; positive regulation of phosphoinositide 3-kinase cascade; positive regulation of protein kinase B signaling cascade; response to ethanol; response to cytokine stimulus; neural tube closure; gene expression; spermatogenesis; positive regulation of transcription from RNA polymerase II promoter; steroid hormone mediated signaling; positive regulation of neuron differentiation; negative regulation of transcription, DNA-dependent; apoptotic cell clearance; negative regulation of apoptosis
Disease: Acute Promyelocytic Leukemia
|NCBI Summary:||This gene represents a nuclear retinoic acid receptor. The encoded protein, retinoic acid receptor alpha, regulates transcription in a ligand-dependent manner. This gene has been implicated in regulation of development, differentiation, apoptosis, granulopoeisis, and transcription of clock genes. Translocations between this locus and several other loci have been associated with acute promyelocytic leukemia. Alternatively spliced transcript variants have been found for this locus.[provided by RefSeq, Sep 2010]|
|NCBI GenInfo Identifier:||133483|
|NCBI Gene ID:||5914|
|UniProt Secondary Accession:||P10276,P78456, Q13440, Q13441, Q96S41, Q9NQS0, B8Y636|
|UniProt Related Accession:||P10276|
|Molecular Weight:||39,700 Da|
|NCBI Full Name:||Retinoic acid receptor alpha|
|NCBI Synonym Full Names:||retinoic acid receptor, alpha|
|NCBI Official Symbol:||RARA|
|NCBI Official Synonym Symbols:||RAR; NR1B1|
|NCBI Protein Information:||retinoic acid receptor alpha; RAR-alpha; retinoic acid receptor, alpha polypeptide; nuclear receptor subfamily 1 group B member 1; retinoic acid nuclear receptor alpha variant 1; retinoic acid nuclear receptor alpha variant 2; nucleophosmin-retinoic acid receptor alpha fusion protein NPM-RAR long form|
|UniProt Protein Name:||Retinoic acid receptor alpha|
|UniProt Synonym Protein Names:||Nuclear receptor subfamily 1 group B member 1|
|Protein Family:||Retinoic acid receptor|
|UniProt Gene Name:||RARA|
|UniProt Entry Name:||RARA_HUMAN|