Mouse RARB / Retinoic acid receptor beta ELISA Kit

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SKU:
MOFI00333
Product Type:
ELISA Kit
Size:
96 Assays
Uniprot:
P22605
Sensitivity:
28.125pg/ml
Range:
46.875-3000pg/ml
ELISA Type:
Sandwich ELISA, Double Antibody
Synonyms:
RarBeta, RARB, Retinoic acid receptor beta, RAR-beta, RAR-epsilon, HBV-activated protein, Nuclear receptor subfamily 1 group B member 2
Reactivity:
Mouse
Frequently bought together:

Description

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Product Name:Mouse RARB / Retinoic acid receptor beta ELISA Kit
Product Code:MOFI00333
Size:96 Assays
Alias:Rarbeta, RARB, Retinoic acid receptor beta, RAR-beta, RAR-epsilon, HBV-activated protein, Nuclear receptor subfamily 1 group B member 2
Detection Method:Sandwich ELISA
Application:This immunoassay kit allows for the in vitro quantitative determination of Mouse Rarbeta concentrations in serum plasma and other biological fluids.
Sensitivity:28.125pg/ml
Range:46.875-3000pg/ml
Storage:4°C for 6 months
Note:For Research Use Only
Recovery:Matrices listed below were spiked with certain level of Mouse Rarbeta and the recovery rates were calculated by comparing the measured value to the expected amount of Mouse Rarbeta in samples.
MatrixRecovery range(%)Average(%)
serum(n=5)91-10296
EDTA plasma(n=5)90-9993
UFH plasma(n=5)89-10296
Linearity:The linearity of the kit was assayed by testing samples spiked with appropriate concentration of Mouse Rarbeta and their serial dilutions. The results were demonstrated by the percentage of calculated concentration to the expected.
Sample1:21:41:8
serum(n=5)86-101%85-105%88-101%
EDTA plasma(n=5)82-100%82-98%84-97%
UFH plasma(n=5)84-99%82-96%80-99%
Intra Assay:CV <8%
Inter Assay:CV <10%
ComponentQuantityStorage
ELISA Microplate (Dismountable)8-12 strips4°C for 6 months
Lyophilized Standard24°C/-20°C
Sample/Standard Dilution Buffer20ml4°C
Biotin-labeled Antibody(Concentrated)120ul4°C (Protect from light)
Antibody Dilution Buffer10ml4°C
HRP-Streptavidin Conjugate(SABC)120ul4°C (Protect from light)
SABC Dilution Buffer10ml4°C
TMB Substrate10ml4°C (Protect from light)
Stop Solution10ml4°C
Wash Buffer(25X)30ml4°C
Plate Sealer5-

Other materials and equipment required:

  • Microplate reader with 450 nm wavelength filter
  • Multichannel Pipette, Pipette, microcentrifuge tubes and disposable pipette tips
  • Incubator
  • Deionized or distilled water
  • Absorbent paper
  • Buffer resevoir
UniprotP22605
UniProt Protein Function:RARB: is a receptor for retinoic acid, a potent mammalian morphogen and teratogen that has profound effects on vertebrate development. RARB is a member of the nuclear receptor superfamily. Controls cell function by directly regulating gene expression. Composed of three domains: a modulating N-terminal domain, a DNA-binding domain and a C-terminal steroid-binding domain. Four splice-variant isoforms have been described. Isoform beta-1 and beta-2 are nuclear and isoform beta-4 cytoplasmic.
UniProt Protein Details:

Protein type:Oncoprotein; Motility/polarity/chemotaxis; Nuclear receptor

Cellular Component: nucleoplasm; nucleus

Molecular Function:DNA binding; drug binding; metal ion binding; protein complex binding; retinoic acid receptor activity; retinoid X receptor binding; sequence-specific DNA binding; steroid hormone receptor activity; transcription factor activity; zinc ion binding

Biological Process: embryonic eye morphogenesis; embryonic gut development; embryonic hindlimb morphogenesis; glandular epithelial cell development; multicellular organism growth; negative regulation of apoptosis; negative regulation of cell proliferation; negative regulation of chondrocyte differentiation; negative regulation of transcription from RNA polymerase II promoter; neurogenesis; positive regulation of apoptosis; positive regulation of cell proliferation; positive regulation of neuron differentiation; positive regulation of programmed cell death; positive regulation of transcription from RNA polymerase II promoter; regulation of myelination; regulation of transcription, DNA-dependent; retina development in camera-type eye; retinoic acid receptor signaling pathway; steroid hormone mediated signaling; striatum development; transcription, DNA-dependent; ureteric bud development; ventricular cardiac muscle cell differentiation

UniProt Code:P22605
NCBI GenInfo Identifier:576795732
NCBI Gene ID:218772
NCBI Accession:NP_001276689.1
UniProt Secondary Accession:P22605,P11417, P22604,
UniProt Related Accession:P22605
Molecular Weight:44,889 Da
NCBI Full Name:retinoic acid receptor beta isoform beta3
NCBI Synonym Full Names:retinoic acid receptor, beta
NCBI Official Symbol:Rarb  
NCBI Official Synonym Symbols:Hap; Nr1b2; A830025K23  
NCBI Protein Information:retinoic acid receptor beta
UniProt Protein Name:Retinoic acid receptor beta
UniProt Synonym Protein Names:Nuclear receptor subfamily 1 group B member 2
Protein Family:Retinoic acid receptor
UniProt Gene Name:Rarb  
UniProt Entry Name:RARB_MOUSE

*Note: Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.

StepProcedure
1. Set standard, test sample and control (zero) wells on the pre-coated plate respectively, and then, record their positions. It is recommended to measure each standard and sample in duplicate. Wash plate 2 times before adding standard, sample and control (zero) wells!
2.Aliquot 0.1ml standard solutions into the standard wells.
3. Add 0.1 ml of Sample / Standard dilution buffer into the control (zero) well.
4. Add 0.1 ml of properly diluted sample (Human serum, plasma, tissue homogenates and other biological fluids.) into test sample wells.
5.Seal the plate with a cover and incubate at 37 °C for 90 min.
6. Remove the cover and discard the plate content, clap the plate on the absorbent filter papers or other absorbent material. Do NOT let the wells completely dry at any time. Wash plate X2.
7. Add 0.1 ml of Biotin- detection antibody working solution into the above wells (standard, test sample & zero wells). Add the solution at the bottom of each well without touching the side wall.
8.Seal the plate with a cover and incubate at 37°C for 60 min.
9. Remove the cover, and wash plate 3 times with Wash buffer. Let wash buffer rest in wells for 1 min between each wash.
10. Add 0.1 ml of SABC working solution into each well, cover the plate and incubate at 37°C for 30 min.
11. Remove the cover and wash plate 5 times with Wash buffer, and each time let the wash buffer stay in the wells for 1-2 min.
12. Add 90 µL of TMB substrate into each well, cover the plate and incubate at 37°C in dark within 10-20 min. (Note: This incubation time is for reference use only, the optimal time should be determined by end user.) And the shades of blue can be seen in the first 3-4 wells (with most concentrated standard solutions), the other wells show no obvious color.
13. Add 50 µL of Stop solution into each well and mix thoroughly. The color changes into yellow immediately.
14. Read the O.D. absorbance at 450 nm in a microplate reader immediately after adding the stop solution.

When carrying out an ELISA assay it is important to prepare your samples in order to achieve the best possible results. Below we have a list of procedures for the preparation of samples for different sample types.

Sample TypeProtocol
Serum:

If using serum separator tubes, allow samples to clot for 30 minutes at room temperature. Centrifuge for 10 minutes at 1,000x g. Collect the serum fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles.

If serum separator tubes are not being used, allow samples to clot overnight at 2-8°C. Centrifuge for 10 minutes at 1,000x g. Remove serum and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles.

Plasma:

Collect plasma using EDTA or heparin as an anticoagulant. Centrifuge samples at 4°C for 15 mins at 1000 - g within 30 mins of collection. Collect the plasma fraction and assay promptly or aliquot and store the samples at -80°C. Avoid multiple freeze-thaw cycles. Note: Over haemolysed samples are not suitable for use with this kit.

Urine & Cerebrospinal Fluid:

Collect the urine (mid-stream) in a sterile container, centrifuge for 20 mins at 2000-3000 rpm. Remove supernatant and assay immediately. If any precipitation is detected, repeat the centrifugation step. A similar protocol can be used for cerebrospinal fluid.

Cell culture supernatant:

Collect the cell culture media by pipette, followed by centrifugation at 4°C for 20 mins at 1500 rpm. Collect the clear supernatant and assay immediately.

Cell lysates:

Solubilize cells in lysis buffer and allow to sit on ice for 30 minutes. Centrifuge tubes at 14,000 x g for 5 minutes to remove insoluble material. Aliquot the supernatant into a new tube and discard the remaining whole cell extract. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20°C.

Tissue homogenates:

The preparation of tissue homogenates will vary depending upon tissue type. Rinse tissue with 1X PBS to remove excess blood & homogenize in 20ml of 1X PBS (including protease inhibitors) and store overnight at ≤ -20°C. Two freeze-thaw cycles are required to break the cell membranes. To further disrupt the cell membranes you can sonicate the samples. Centrifuge homogenates for 5 mins at 5000xg. Remove the supernatant and assay immediately or aliquot and store at -20°C or -80°C.

Tissue lysates:

Rinse tissue with PBS, cut into 1-2 mm pieces, and homogenize with a tissue homogenizer in PBS. Add an equal volume of RIPA buffer containing protease inhibitors and lyse tissues at room temperature for 30 minutes with gentle agitation. Centrifuge to remove debris. Quantify total protein concentration using a total protein assay. Assay immediately or aliquot and store at ≤ -20 °C.

Breast Milk:

Collect milk samples and centrifuge at 10,000 x g for 60 min at 4°C. Aliquot the supernatant and assay. For long term use, store samples at -80°C. Minimize freeze/thaw cycles.

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