Human GPRC5A (G Protein Coupled Receptor, Family C, Group 5, Member A) ELISA Kit (HUES02180)

SKU:
HUES02180
€539

Description

ELISA Kit Technical ManualMSDS

Human GPRC5A (G Protein Coupled Receptor, Family C, Group 5, Member A) 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 GPRC5A . 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 GPRC5A 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 GPRC5A, 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 GPRC5A. The concentration of Human GPRC5A in samples can be calculated by comparing the OD of the samples to the standard curve.

Assay typeSandwich
Format96T
Assay time4.5h
ReactivityHuman
Detection MethodColormetric
Detection Range0.31-20 ng/mL
Sensitivity0.19 ng/mL
Sample Volume Required Per Well100uL
Sample TypeSerum, plasma and other biological fluids

Specificity

This kit recognizes Human GPRC5A in samples. No significant cross-reactivity or interference between Human GPRC5A and analogues was observed.

Typical Data

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.

Concentration
(ng/mL)
O.D Average Corrected
202.406
2.41
2.4082.322
101.623
1.625
1.6241.538
50.925
0.923
0.9240.838
2.50.428
0.46
0.4440.358
1.250.261
0.243
0.2520.166
0.630.191
0.179
0.1850.099
0.310.131
0.141
0.1360.05
00.085
0.087
0.086--

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human GPRC5A were tested 20 times on one plate, respectively.

Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human GPRC5A were tested on 3 different plates, 20 replicates in each plate.

Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (ng/mL)0.963.218.061.013.368.27
Standard deviation0.060.150.360.060.190.37
C V (%)6.254.674.475.945.654.47

Recovery

The recovery of Human GPRC5A spiked at three different levels in samples throughout the range of the assay was evaluated in various matrices.

Sample Type Range (%) Average Recovery (%)
Serum (n=5)95-106101
EDTA plasma (n=5)90-10395
Cell culture media (n=5)90-10396

Linearity

Samples were spiked with high concentrations of Human GPRC5A 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)
1:2Range (%)89-10089-9988-104
Average (%)949495
1:4Range (%)94-10979-9385-97
Average (%)1008591
1:8Range (%)86-9983-9885-96
Average (%)939091
1:16Range (%)91-10579-9287-101
Average (%)988694

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.

Item Specifications Storage
Micro ELISA Plate(Dismountable)8 wells X12 strips-20'C, 6 months
Reference Standard2 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 Diluent1 vial, 20 mL4'C, 6 months
Biotinylated Detection Ab Diluent1 vial, 14 mL
HRP Conjugate Diluent1 vial, 14 mL
Concentrated Wash Buffer (25X)1 vial, 30 mL
Substrate Reagent1 vial, 10 mL4'C(shading light)
Stop Solution1 vial, 10 mL4'C
Plate Sealer5 pieces
Product Description1 copy
Certificate of Analysis1 copy

Human GPRC5A (G Protein Coupled Receptor, Family C, Group 5, Member A) ELISA Kit (HUES02180) 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 100 µL of standard solutions into the standard wells.
    3. Add 100 µL of Sample / Standard dilution buffer into the control (zero) well.
    4. Add 100 µL 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 100 µL 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 350 µL 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 100 µL 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 90 µL 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 GPRC5A (G Protein Coupled Receptor, Family C, Group 5, Member A) ELISA Kit (HUES02180) Protein Information

UniProt Protein Function:RAIG1: a G protein coupled receptor that is induced by all-trans retinoic acid (ATRA). Retinoic acids (RA) are a group of pleiotropic signaling molecules that regulate a broad range of physiologic and developmental processes. RAIG1 is expressed in lung tissue and in several lung cancer cell lines. This retinoic acid-inducible GPCR provides evidence for an interaction between retinoid and G-protein signaling pathways.
UniProt Protein Details:

Protein type:Membrane protein, integral; GPCR, family 3; Membrane protein, multi-pass; Receptor, GPCR

Chromosomal Location of Human Ortholog: 12p13.1

Cellular Component: cytoplasmic vesicle membrane; integral to plasma membrane; intracellular membrane-bound organelle; plasma membrane

Molecular Function:G-protein coupled receptor activity; protein binding

Biological Process: G-protein coupled receptor protein signaling pathway; negative regulation of epidermal growth factor receptor activity; signal transduction

NCBI Summary:This gene encodes a member of the type 3 G protein-coupling receptor family, characterized by the signature 7-transmembrane domain motif. The encoded protein may be involved in interaction between retinoid acid and G protein signalling pathways. Retinoic acid plays a critical role in development, cellular growth, and differentiation. This gene may play a role in embryonic development and epithelial cell differentiation. [provided by RefSeq, Jul 2008]
UniProt Code:Q8NFJ5
NCBI GenInfo Identifier:46396943
NCBI Gene ID:9052
NCBI Accession:Q8NFJ5.2
UniProt Secondary Accession:Q8NFJ5,O95357, B3KV45,
UniProt Related Accession:Q8NFJ5
Molecular Weight:40,251 Da
NCBI Full Name:Retinoic acid-induced protein 3
NCBI Synonym Full Names:G protein-coupled receptor class C group 5 member A
NCBI Official Symbol:GPRC5A
NCBI Official Synonym Symbols:RAI3; TIG1; RAIG1; GPCR5A; PEIG-1
NCBI Protein Information:retinoic acid-induced protein 3
UniProt Protein Name:Retinoic acid-induced protein 3
UniProt Synonym Protein Names:G-protein coupled receptor family C group 5 member A; Phorbol ester induced gene 1
Protein Family:Retinoic acid-induced protein
UniProt Gene Name:GPRC5A
UniProt Entry Name:RAI3_HUMAN
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Additional Information

Product type:
ELISA
Reactivity:
Human
ELISA Type:
Sandwich
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