|Detection range:||62.50-4000 fmol/mL|
|Sample type:||Serum, plasma and other biological fluids|
|Repeatability:||CV < 15%|
|Specificity:||This kit recognizes Rat CREB in samples. No significant cross-reactivity or interference between Rat CREB 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 CREB. 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 CREB 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 CREB, 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 CREB. The concentration of Rat CREB in the samples can be calculated by comparing the RLU of the samples to the standard curve.
|UniProt Protein Function:||CREB: a transcription factor of the leucine zipper family of DNA binding proteins. Binds as a homodimer to the cAMP-responsive element (CRE). Phosphorylated by several protein kinases, and induces transcription of genes in response to hormonal stimulation of the cAMP pathway. Two splice-variant isoforms have been described.|
|UniProt Protein Details:|
Protein type:DNA-binding; Transcription factor
Cellular Component: axon; chromatin; mitochondrial matrix; mitochondrion; nuclear chromatin; nucleoplasm; nucleus; transcription factor complex
Molecular Function:DNA binding; double-stranded DNA binding; enzyme binding; histone acetyltransferase binding; Hsp70 protein binding; identical protein binding; protein binding; RNA polymerase II transcription factor activity, enhancer binding; sequence-specific DNA binding; transcription factor activity; transcription factor binding
Biological Process: aging; axonogenesis; circadian rhythm; lactation; mammary gland development; memory; pituitary gland development; positive regulation of apoptosis; positive regulation of fat cell differentiation; positive regulation of hormone secretion; positive regulation of lipid biosynthetic process; positive regulation of multicellular organism growth; positive regulation of osteoclast differentiation; positive regulation of transcription from RNA polymerase II promoter; positive regulation of transcription, DNA-dependent; positive regulation of transcriptional preinitiation complex assembly; positive regulation of transforming growth factor-beta3 production; protein amino acid phosphorylation; protein stabilization; regulation of apoptosis; regulation of cell size; regulation of circadian rhythm; regulation of fibroblast proliferation; regulation of transcription, DNA-dependent; response to activity; response to drug; response to glucagon stimulus; response to hypoxia; response to nicotine; response to organic substance; secretory granule organization and biogenesis; transcription from RNA polymerase II promoter; transforming growth factor beta receptor signaling pathway; visual learning
|NCBI Summary:||binds the cAMP response element in many gene promoters and regulates transcription [RGD, Feb 2006]|
|NCBI GenInfo Identifier:||117435|
|NCBI Gene ID:||81646|
|NCBI Accession:||P15337. 1|
|UniProt Related Accession:||P15337|
|Molecular Weight:||35,081 Da|
|NCBI Full Name:||Cyclic AMP-responsive element-binding protein 1|
|NCBI Synonym Full Names:||cAMP responsive element binding protein 1|
|NCBI Official Symbol:||Creb1|
|NCBI Official Synonym Symbols:||Creb|
|NCBI Protein Information:||cyclic AMP-responsive element-binding protein 1|
|UniProt Protein Name:||Cyclic AMP-responsive element-binding protein 1|
|Protein Family:||Cyclic AMP-responsive element-binding protein|
|UniProt Gene Name:||Creb1|
|UniProt Entry Name:||CREB1_RAT|
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.
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Rat CREB were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Rat CREB were tested on 3 different plates, 20 replicates in each plate.
|Intra-assay Precision||Inter-assay Precision|
|C V (%)||12.83||8.93||10.36||12.27||8.52||10.19|
The recovery of Rat CREB 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)||99-114||106|
|Cell culture media (n=5)||97-108||103|
Samples were spiked with high concentrations of Rat CREB 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)|
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 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.