Rat Signaling ELISA Kits 2
Rat TGF- beta1(Transforming Growth Factor beta1) CLIA Kit (RTES00052)
- SKU:
- RTES00052
- Product Type:
- ELISA Kit
- ELISA Type:
- CLIA Kit
- Size:
- 96 Assays
- Sensitivity:
- 7.5pg/mL
- Range:
- 12.5-800pg/mL
- ELISA Type:
- Sandwich
- Synonyms:
- TGF-B1, TGFB, TGFbeta, CED, DPD1, LAP
- 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: | 12.50-800 pg/mL |
| Sensitivity: | 7.50 pg/mL |
| Sample volume: | 100µL |
| Sample type: | Serum, plasma and other biological fluids |
| Repeatability: | CV < 15% |
| Specificity: | This kit recognizes Rat TGF- beta1 in samples. No significant cross-reactivity or interference between Rat TGF- beta1 and analogues was observed. |
This CLIA 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 TGF- beta1. Standards or samples are added to the micro CLIA plate wells and combined with the specific antibody. Then a biotinylated detection antibody specific for Rat TGF- beta1 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 TGF- beta1, 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 TGF- beta1. You can calculate the concentration of Rat TGF- beta1 in the samples by comparing the RLU value of the samples to the standard curve.
| UniProt Protein Function: | TGFB1: Multifunctional protein that controls proliferation, differentiation and other functions in many cell types. Many cells synthesize TGFB1 and have specific receptors for it. It positively and negatively regulates many other growth factors. It plays an important role in bone remodeling as it is a potent stimulator of osteoblastic bone formation, causing chemotaxis, proliferation and differentiation in committed osteoblasts. Homodimer; disulfide-linked, or heterodimer with TGFB2. Secreted and stored as a biologically inactive form in the extracellular matrix in a 290 kDa complex (large latent TGF-beta1 complex) containing the TGFB1 homodimer, the latency-associated peptide (LAP), and the latent TGFB1 binding protein-1 (LTBP1). The complex without LTBP1 is known as the'small latent TGF-beta1 complex'. Dissociation of the TGFB1 from LAP is required for growth factor activation and biological activity. Release of the large latent TGF-beta1 complex from the extracellular matrix is carried out by the matrix metalloproteinase MMP3. May interact with THSD4; this interaction may lead to sequestration by FBN1 microfibril assembly and attenuation of TGFB signaling. Interacts with the serine proteases, HTRA1 and HTRA3: the interaction with either inhibits TGFB1-mediated signaling. The HTRA protease activity is required for this inhibition. Interacts with CD109, DPT and ASPN. Activated in vitro at pH below 3. 5 and over 12. 5. Highly expressed in bone. Abundantly expressed in articular cartilage and chondrocytes and is increased in osteoarthritis (OA). Co-localizes with ASPN in chondrocytes within OA lesions of articular cartilage. Belongs to the TGF-beta family. |
| UniProt Protein Details: | Protein type:Motility/polarity/chemotaxis; Secreted; Secreted, signal peptide Cellular Component: axon; cell soma; cell surface; cytoplasm; extracellular matrix; extracellular space; microvillus; nucleus; proteinaceous extracellular matrix; secretory granule Molecular Function:antigen binding; cytokine activity; enzyme binding; glycoprotein binding; protein heterodimerization activity; protein homodimerization activity; protein N-terminus binding; protein serine/threonine kinase activator activity; punt binding; transforming growth factor beta receptor binding Biological Process: activation of NF-kappaB transcription factor; adaptive immune response based on somatic recombination of immune receptors built from immunoglobulin superfamily domains; aging; ATP biosynthetic process; cell activation; cell cycle arrest; cell migration; cell proliferation; cellular calcium ion homeostasis; chondrocyte differentiation; common-partner SMAD protein phosphorylation; defense response to fungus, incompatible interaction; endoderm development; epidermal growth factor receptor signaling pathway; epithelial to mesenchymal transition; evasion of host defenses by virus; female pregnancy; germ cell migration; gut development; heart development; hemopoietic progenitor cell differentiation; hyaluronan catabolic process; inflammatory response; inner ear development; intercellular junction assembly and maintenance; lipopolysaccharide-mediated signaling pathway; lung development; lymph node development; mammary gland development; MAPKKK cascade; membrane protein intracellular domain proteolysis; mitotic cell cycle checkpoint; mononuclear cell proliferation; morphogenesis of a branching structure; myelination; myeloid dendritic cell differentiation; negative regulation of blood vessel endothelial cell migration; negative regulation of cell cycle; negative regulation of cell differentiation; negative regulation of cell growth; negative regulation of cell proliferation; negative regulation of cell-cell adhesion; negative regulation of DNA replication; negative regulation of epithelial cell proliferation; negative regulation of fat cell differentiation; negative regulation of immune response; negative regulation of interleukin-17 production; negative regulation of mitotic cell cycle; negative regulation of myoblast differentiation; negative regulation of neuroblast proliferation; negative regulation of ossification; negative regulation of phagocytosis; negative regulation of protein amino acid phosphorylation; negative regulation of release of sequestered calcium ion into cytosol; negative regulation of skeletal muscle development; negative regulation of T cell activation; negative regulation of T cell proliferation; negative regulation of transcription from RNA polymerase II promoter; negative regulation of transcription, DNA-dependent; neural tube closure; neural tube development; Notch signaling pathway; oligodendrocyte development; organ regeneration; phosphate metabolic process; positive regulation of apoptosis; positive regulation of blood vessel endothelial cell migration; positive regulation of bone mineralization; positive regulation of cell migration; positive regulation of cell proliferation; positive regulation of cellular protein metabolic process; positive regulation of chemotaxis; positive regulation of collagen biosynthetic process; positive regulation of epithelial cell proliferation; positive regulation of exit from mitosis; positive regulation of fibroblast proliferation; positive regulation of histone acetylation; positive regulation of histone deacetylation; positive regulation of interleukin-17 production; positive regulation of isotype switching to IgA isotypes; positive regulation of MAP kinase activity; positive regulation of odontogenesis; positive regulation of peptidyl-serine phosphorylation; positive regulation of peptidyl-tyrosine phosphorylation; positive regulation of phosphoinositide 3-kinase activity; positive regulation of protein amino acid dephosphorylation; positive regulation of protein amino acid phosphorylation; positive regulation of protein complex assembly; positive regulation of protein import into nucleus; positive regulation of protein kinase B signaling cascade; positive regulation of protein secretion; positive regulation of regulatory T cell differentiation; positive regulation of smooth muscle cell differentiation; positive regulation of superoxide release; positive regulation of transcription from RNA polymerase II promoter; positive regulation of transcription, DNA-dependent; positive regulation of vascular permeability; protein amino acid phosphorylation; protein export from nucleus; protein import into nucleus, translocation; protein kinase B signaling cascade; receptor catabolic process; regulation of binding; regulation of cell growth; regulation of cell proliferation; regulation of DNA binding; regulation of gene expression; regulation of interleukin-23 production; regulation of MAPKKK cascade; regulation of protein import into nucleus; regulation of regulatory T cell differentiation; regulation of sodium ion transport; regulation of striated muscle development; regulation of transforming growth factor beta receptor signaling pathway; regulatory T cell differentiation; response to drug; response to estradiol stimulus; response to glucose stimulus; response to hypoxia; response to organic cyclic substance; response to organic substance; response to progesterone stimulus; response to radiation; response to vitamin D; response to wounding; salivary gland morphogenesis; SMAD protein complex assembly; SMAD protein nuclear translocation; T cell activation; T cell differentiation; T cell homeostasis; tolerance induction to self antigen; transforming growth factor beta receptor signaling pathway; ureteric bud development; vasculogenesis; ventricular cardiac muscle morphogenesis; wound healing |
| NCBI Summary: | binds the TGFbeta receptor; plays a role in regulation of cell growth and proliferation; induces synthesis of extracellular matrix proteins and may play a role in fibrosis [RGD, Feb 2006] |
| UniProt Code: | P17246 |
| NCBI GenInfo Identifier: | 135677 |
| NCBI Gene ID: | 59086 |
| NCBI Accession: | P17246. 1 |
| UniProt Secondary Accession: | P17246,Q53YM8, |
| UniProt Related Accession: | P17246 |
| Molecular Weight: | 44,329 Da |
| NCBI Full Name: | Transforming growth factor beta-1 |
| NCBI Synonym Full Names: | transforming growth factor, beta 1 |
| NCBI Official Symbol: | Tgfb1Ă‚Â Ă‚Â |
| NCBI Official Synonym Symbols: | Tgfb  |
| NCBI Protein Information: | transforming growth factor beta-1; TGF-beta-1; transforming growth factor, beta-1 |
| UniProt Protein Name: | Transforming growth factor beta-1 |
| UniProt Synonym Protein Names: | |
| Protein Family: | Transforming growth factor |
| UniProt Gene Name: | Tgfb1Ă‚Â Ă‚Â |
| UniProt Entry Name: | TGFB1_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.
| Concentration (pg/mL) | RLU | Average | Corrected |
| 800 | 47980 57304 | 52642 | 52616 |
| 400 | 19812 23656 | 21734 | 21708 |
| 200 | 9977 9439 | 9708 | 9682 |
| 100 | 4545 4559 | 4552 | 4526 |
| 50 | 2261 2115 | 2188 | 2162 |
| 25 | 1112 1008 | 1060 | 1034 |
| 12.50 | 500 518 | 509 | 483 |
| 0 | 26 26 | 26 | -- |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Rat TGF- beta1 were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Rat TGF- beta1 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) | 40.15 | 87.21 | 352.61 | 41.27 | 88.49 | 353.17 |
| Standard deviation | 3.94 | 7.86 | 40.23 | 5.11 | 10.13 | 39.94 |
| C V (%) | 9.81 | 9.01 | 11.41 | 12.38 | 11.45 | 11.31 |
Recovery
The recovery of Rat TGF- beta1 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) | 94-110 | 102 |
| EDTA plasma (n=5) | 87-102 | 94 |
| Cell culture media (n=5) | 94-108 | 102 |
Linearity
Samples were spiked with high concentrations of Rat TGF- beta1 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 (%) | 101-117 | 99-114 | 91-105 |
| Average (%) | 109 | 106 | 97 | |
| 1:4 | Range (%) | 91-105 | 92-105 | 85-97 |
| Average (%) | 99 | 98 | 92 | |
| 1:8 | Range (%) | 92-108 | 94-106 | 102-115 |
| Average (%) | 98 | 101 | 108 | |
| 1:16 | Range (%) | 86-98 | 101-116 | 86-98 |
| Average (%) | 91 | 108 | 93 |
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 Ă—12 strips | -20°C, 6 months |
| Reference Standard | 2 vials | |
| Concentrated Biotinylated Detection Ab (100Ă—) | 1 vial, 120 µL | |
| Concentrated HRP Conjugate (100Ă—) | 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 (25Ă—) | 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 100ĂƒÂ‚µl of standard solutions into the standard wells.
- Add 100ĂƒÂ‚µl of Sample / Standard dilution buffer into the control (zero) well.
- Add 100ĂƒÂ‚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 37ĂƒÂ‚°C.
- Aspirate the liquid from each well, do not wash. Immediately add 100ĂƒÂ‚µL of BiotinylatedDetection Ab working solution to each well. Cover the plate with a plate seal and gently mix. Incubate for 1 hour at 37ĂƒÂ‚°C.
- Aspirate or decant the solution from the plate and add 350ĂƒÂ‚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 100ĂƒÂ‚µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37ĂƒÂ‚°C.
- Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
- Add 100ĂƒÂ‚µL of Substrate mixture solution to each well. Cover with a new plate seal andincubate for no more than 5 min at 37ĂƒÂ‚°C. Protect the plate from light.
- Determine the RLU value of each well immediately.
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