Human RGS4 (Regulator of G Protein Signaling 4) ELISA Kit (HUES02015)
- SKU:
- HUES02015
- Product Type:
- ELISA Kit
- Size:
- 96 Assays
- Uniprot:
- P49798
- Sensitivity:
- 0.09ng/mL
- Range:
- 0.16-10ng/mL
- ELISA Type:
- Sandwich
- Reactivity:
- Human
- Tested Sample Types:
- Serum, plasma and other biological fluids
Description
| Assay type: | Sandwich |
| Format: | 96T |
| Assay time: | 4.5h |
| Reactivity: | Human |
| Detection Method: | Colormetric |
| Detection Range: | 0.16-10 ng/mL |
| Sensitivity: | 0.10 ng/mL |
| Sample Volume Required Per Well: | 100µL |
| Sample Type: | Serum, plasma and other biological fluids |
| Specificity: | This kit recognizes Human RGS4 in samples. No significant cross-reactivity or interference between Human RGS4 and analogues was observed. |
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 RGS4. 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 RGS4 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 RGS4, 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 RGS4. The concentration of Human RGS4 in samples can be calculated by comparing the OD of the samples to the standard curve.
| UniProt Protein Function: | RGS4: Inhibits signal transduction by increasing the GTPase activity of G protein alpha subunits thereby driving them into their inactive GDP-bound form. Activity on G(z)-alpha is inhibited by phosphorylation of the G-protein. Activity on G(z)-alpha and G(i)-alpha-1 is inhibited by palmitoylation of the G-protein. Expressed in brain and heart. Expressed in brain at protein level. Expressed in prefontal and visual cortex. Isoform 4 and isoform 5 are expressed ubiquitously. Isoform 1, isoform 2 and isoform 3 are not expressed in the cerebellum. 5 isoforms of the human protein are produced by alternative splicing. |
| UniProt Protein Details: | Protein type:GAPs, RGS; GAPs Chromosomal Location of Human Ortholog: 1q23. 3 Cellular Component: cytoplasm; nucleus; plasma membrane Molecular Function:calmodulin binding Biological Process: inactivation of MAPK activity; regulation of G-protein coupled receptor protein signaling pathway Disease: Schizophrenia 9 |
| NCBI Summary: | Regulator of G protein signaling (RGS) family members are regulatory molecules that act as GTPase activating proteins (GAPs) for G alpha subunits of heterotrimeric G proteins. RGS proteins are able to deactivate G protein subunits of the Gi alpha, Go alpha and Gq alpha subtypes. They drive G proteins into their inactive GDP-bound forms. Regulator of G protein signaling 4 belongs to this family. All RGS proteins share a conserved 120-amino acid sequence termed the RGS domain. Regulator of G protein signaling 4 protein is 37% identical to RGS1 and 97% identical to rat Rgs4. This protein negatively regulate signaling upstream or at the level of the heterotrimeric G protein and is localized in the cytoplasm. Alternatively spliced transcript variants have been found for this gene. [provided by RefSeq, Jul 2008] |
| UniProt Code: | P49798 |
| NCBI GenInfo Identifier: | 1710146 |
| NCBI Gene ID: | 5999 |
| NCBI Accession: | P49798. 1 |
| UniProt Secondary Accession: | P49798,A7XA56, A7XA58, A7XA59, A7YVV7, B1APZ3, |
| UniProt Related Accession: | P49798 |
| Molecular Weight: | 21,452 Da |
| NCBI Full Name: | Regulator of G-protein signaling 4 |
| NCBI Synonym Full Names: | regulator of G-protein signaling 4 |
| NCBI Official Symbol: | RGS4 |
| NCBI Official Synonym Symbols: | RGP4; SCZD9 |
| NCBI Protein Information: | regulator of G-protein signaling 4 |
| UniProt Protein Name: | Regulator of G-protein signaling 4 |
| Protein Family: | Regulator of G-protein signaling |
| UniProt Gene Name: | RGS4 |
| UniProt Entry Name: | RGS4_HUMAN |
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 |
| 10 | 2.322 2.366 | 2.344 | 2.278 |
| 5 | 1.515 1.555 | 1.535 | 1.469 |
| 2.5 | 0.842 0.834 | 0.838 | 0.772 |
| 1.25 | 0.375 0.393 | 0.384 | 0.318 |
| 0.63 | 0.254 0.254 | 0.254 | 0.188 |
| 0.32 | 0.165 0.151 | 0.158 | 0.092 |
| 0.16 | 0.109 0.119 | 0.114 | 0.048 |
| 0 | 0.064 0.068 | 0.066 | -- |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human RGS4 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human RGS4 were tested on 3 different plates, 20 replicates in each plate.
| Intra-assay Precision | Inter-assay Precision | |||||
| Sample | 1 | 2 | 3 | 1 | 2 | 3 |
| n | 20 | 20 | 20 | 20 | 20 | 20 |
| Mean (ng/mL) | 0.52 | 1.52 | 4.66 | 0.47 | 1.51 | 5.10 |
| Standard deviation | 0.03 | 0.08 | 0.25 | 0.03 | 0.08 | 0.26 |
| C V (%) | 5.77 | 5.26 | 5.36 | 6.38 | 5.30 | 5.10 |
Recovery
The recovery of Human RGS4 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) | 95-110 | 102 |
| EDTA plasma (n=5) | 94-106 | 100 |
| Cell culture media (n=5) | 94-109 | 101 |
Linearity
Samples were spiked with high concentrations of Human RGS4 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) | ||
| 1:2 | Range (%) | 85-95 | 95-112 | 90-104 |
| Average (%) | 90 | 103 | 96 | |
| 1:4 | Range (%) | 92-109 | 80-90 | 85-98 |
| Average (%) | 99 | 86 | 90 | |
| 1:8 | Range (%) | 92-106 | 87-101 | 85-97 |
| Average (%) | 97 | 93 | 92 | |
| 1:16 | Range (%) | 93-108 | 83-98 | 82-95 |
| Average (%) | 99 | 90 | 87 |
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 ×12 strips | -20°C, 6 months |
| Reference Standard | 2 vials | |
| Concentrated Biotinylated Detection Ab (100×) | 1 vial, 120 µL | |
| Concentrated HRP Conjugate (100×) | 1 vial, 120 µL | -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 (25×) | 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 |
- 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.
- Aliquot 100 µL of standard solutions into the standard wells.
- Add 100 µL of Sample / Standard dilution buffer into the control (zero) well.
- Add 100 µL of properly diluted sample (serum, plasma, tissue homogenates and otherbiological fluids) into test sample wells.
- Cover the plate with the sealer provided in the kit and incubate for 90 min at 37 °C.
- 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.
- 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.
- Add 100 µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37 °C.
- Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
- 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.
- Add 50 µL of Stop Solution to each well. Note: Adding the stop solution should be done inthe same order as the substrate solution.
- Determine the optical density (OD value) of each well immediately with a microplate readerset at 450 nm.
