Rat Signaling ELISA Kits 3
Rat GRB10 (Growth Factor Receptor Bound Protein 10) CLIA Kit (RTES00270)
- SKU:
- RTES00270
- Product Type:
- ELISA Kit
- ELISA Type:
- CLIA Kit
- Size:
- 96 Assays
- Sensitivity:
- 18.75pg/mL
- Range:
- 31.25-2000pg/mL
- ELISA Type:
- Sandwich
- Reactivity:
- Rat
- Sample Type:
- Serum, plasma and other biological fluids
Description
| Assay type: | Sandwich |
| Format: | 96T |
| Assay time: | 4.5h |
| Reactivity: | Rat |
| Detection method: | Chemiluminescence |
| Detection range: | 31.25-2000 pg/mL |
| Sensitivity: | 18.75 pg/mL |
| Sample volume: | 100µL |
| Sample type: | Serum, plasma and other biological fluids |
| Repeatability: | CV < 15% |
| Specificity: | This kit recognizes Rat GRB10 in samples. No significant cross-reactivity or interference between Rat GRB10 and analogues was observed. |
This kit uses Sandwich-CLIA as the method. The micro CLIA plate provided in this kit has been pre-coated with an antibody specific to Rat GRB10. Standards or samples are added to the appropriate micro CLIA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Rat GRB10 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 Rat GRB10, biotinylated detection antibody and Avidin-HRP conjugate will appear fluorescence. The Relative light unit (RLU) value is measured spectrophotometrically by the Chemiluminescence immunoassay analyzer. The RLU value is positively associated with the concentration of Rat GRB10. The concentration of Rat GRB10 in the samples can be calculated by comparing the RLU of the samples to the standard curve.
| UniProt Protein Function: | Adapter protein which modulates coupling of a number of cell surface receptor kinases with specific signaling pathways. Binds to, and suppress signals from, activated receptors tyrosine kinases, including the insulin (INSR) and insulin-like growth factor (IGF1R) receptors. The inhibitory effect can be achieved by 2 mechanisms: interference with the signaling pathway and increased receptor degradation. Delays and reduces AKT1 phosphorylation in response to insulin stimulation. Blocks association between INSR and IRS1 and IRS2 and prevents insulin-stimulated IRS1 and IRS2 tyrosine phosphorylation. Recruits NEDD4 to IGF1R, leading to IGF1R ubiquitination, increased internalization and degradation by both the proteasomal and lysosomal pathways. A similar role in the mediation of ubiquitination has also been suggested with INSR. Negatively regulates Wnt signaling by interacting with LRP6 intracellular portion and interfering with the binding of AXIN1 to LRP6. Positive regulator of the KDR/VEGFR-2 signaling pathway. May inhibit NEDD4-mediated degradation of KDR/VEGFR-2. |
| UniProt Code: | P0CE43 |
| NCBI GenInfo Identifier: | 1046859290 |
| NCBI Gene ID: | 498416 |
| NCBI Accession: | XP_017454822. 1 |
| UniProt Related Accession: | P0CE43 |
| Molecular Weight: | 67,882 Da |
| NCBI Full Name: | growth factor receptor-bound protein 10 isoform X2 |
| NCBI Synonym Full Names: | growth factor receptor bound protein 10 |
| NCBI Official Symbol: | Grb10 |
| NCBI Official Synonym Symbols: | RGD1566234 |
| NCBI Protein Information: | growth factor receptor-bound protein 10 |
| UniProt Protein Name: | Growth factor receptor-bound protein 10 |
| UniProt Synonym Protein Names: | GRB10 adapter protein |
| Protein Family: | Growth factor receptor-bound protein |
| UniProt Gene Name: | Grb10 |
As the RLU 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 (pg/mL) | RLU | Average | Corrected |
| 2000 | 53200 58242 | 55721 | 55693 |
| 1000 | 23231 24717 | 23974 | 23946 |
| 500 | 11457 10607 | 11032 | 11004 |
| 250 | 5045 5543 | 5294 | 5266 |
| 125 | 2787 2429 | 2608 | 2580 |
| 62.5 | 1352 1270 | 1311 | 1283 |
| 31.25 | 652 696 | 674 | 646 |
| 0 | 27 29 | 28 | -- |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Rat GRB10 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Rat GRB10 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 (pg/mL) | 96.02 | 297.65 | 717.77 | 89.02 | 273.79 | 746.53 |
| Standard deviation | 8.97 | 31.22 | 73.86 | 9.33 | 23.22 | 74.35 |
| C V (%) | 9.34 | 10.49 | 10.29 | 10.48 | 8.48 | 9.96 |
Recovery
The recovery of Rat GRB10 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) | 98-111 | 104 |
| EDTA plasma (n=5) | 92-105 | 99 |
| Cell culture media (n=5) | 93-107 | 99 |
Linearity
Samples were spiked with high concentrations of Rat GRB10 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 (%) | 97-113 | 96-110 | 93-110 |
| Average (%) | 104 | 103 | 100 | |
| 1:4 | Range (%) | 100-118 | 89-100 | 96-112 |
| Average (%) | 108 | 95 | 102 | |
| 1:8 | Range (%) | 88-102 | 94-111 | 92-104 |
| Average (%) | 94 | 102 | 98 | |
| 1:16 | Range (%) | 92-106 | 91-104 | 84-98 |
| Average (%) | 99 | 98 | 90 |
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 CLIA 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 A | 1 vial, 5 mL | 4°C (shading light) |
| Substrate Reagent B | 1 vial, 5 mL | 4°C (shading light) |
| 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 100µl of standard solutions into the standard wells.
- Add 100µl of Sample / Standard dilution buffer into the control (zero) well.
- Add 100µl 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 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.
- 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.
- Add 100µL 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 100µL of Substrate mixture solution to each well. Cover with a new plate seal andincubate for no more than 5 min at 37°C. Protect the plate from light.
- Determine the RLU value of each well immediately.
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