Human Cell Biology ELISA Kits 1
Human PDGFB (Platelet Derived Growth Factor Subunit B) ELISA Kit (HUES02669)
- SKU:
- HUES02669
- Product Type:
- ELISA Kit
- Size:
- 96 Assays
- Uniprot:
- P01127
- Sensitivity:
- 14.06pg/mL
- Range:
- 23.44-1500pg/mL
- ELISA Type:
- Sandwich
- Synonyms:
- PDGF-2, PDGF2, SIS, SSV, c-sis
- Reactivity:
- Human
- Sample Type:
- Serum, plasma and other biological fluids
- Research Area:
- Cell Biology
Description
| Assay type: | Sandwich |
| Format: | 96T |
| Assay time: | 4.5h |
| Reactivity: | Human |
| Detection Method: | Colormetric |
| Detection Range: | 23.44-1500 pg/mL |
| Sensitivity: | 14.06 pg/mL |
| Sample Volume Required Per Well: | 100µL |
| Sample Type: | Serum, plasma and other biological fluids |
| Specificity: | This kit recognizes Human PDGFB in samples. No significant cross-reactivity or interference between Human PDGFB 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 PDGFB. 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 PDGFB 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 PDGFB, 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 PDGFB. The concentration of Human PDGFB in samples can be calculated by comparing the OD of the samples to the standard curve.
| UniProt Protein Function: | PDGFB: Growth factor that plays an essential role in the regulation of embryonic development, cell proliferation, cell migration, survival and chemotaxis. Potent mitogen for cells of mesenchymal origin. Required for normal proliferation and recruitment of pericytes and vascular smooth muscle cells in the central nervous system, skin, lung, heart and placenta. Required for normal blood vessel development, and for normal development of kidney glomeruli. Plays an important role in wound healing. Signaling is modulated by the formation of heterodimers with PDGFA. A chromosomal aberration involving PDGFB is found in dermatofibrosarcoma protuberans. Translocation t(17;22)(q22;q13) with PDGFB. Belongs to the PDGF/VEGF growth factor family. |
| UniProt Protein Details: | Protein type:Oncoprotein; Secreted, signal peptide; Motility/polarity/chemotaxis; Secreted Chromosomal Location of Human Ortholog: 22q13. 1 Cellular Component: Golgi membrane; extracellular space; cell surface; basolateral plasma membrane; endoplasmic reticulum lumen; cytoplasm; extracellular region Molecular Function:collagen binding; identical protein binding; protein binding; protein homodimerization activity; growth factor activity; protein heterodimerization activity; platelet-derived growth factor binding; platelet-derived growth factor receptor binding; superoxide-generating NADPH oxidase activator activity; chemoattractant activity Biological Process: extracellular matrix organization and biogenesis; positive regulation of cyclin-dependent protein kinase activity; nerve growth factor receptor signaling pathway; positive regulation of transcription, DNA-dependent; heart development; positive regulation of fibroblast growth factor receptor signaling pathway; cell projection biogenesis; protein amino acid phosphorylation; positive regulation of MAP kinase activity; monocyte chemotaxis; positive regulation of fibroblast proliferation; positive chemotaxis; transforming growth factor beta receptor signaling pathway; cell growth; positive regulation of mitotic cell cycle, embryonic; response to drug; substrate-bound cell migration; platelet activation; fibroblast growth factor receptor signaling pathway; positive regulation of mitosis; positive regulation of chemotaxis; activation of protein kinase B; positive regulation of blood vessel endothelial cell migration; positive regulation of protein amino acid autophosphorylation; positive regulation of phosphoinositide 3-kinase cascade; positive regulation of peptidyl-tyrosine phosphorylation; activation of protein kinase activity; blood vessel morphogenesis; positive regulation of endothelial cell proliferation; negative regulation of transcription, DNA-dependent; actin cytoskeleton organization and biogenesis; embryonic placenta development; peptidyl-tyrosine phosphorylation; positive regulation of smooth muscle cell proliferation; platelet-derived growth factor receptor signaling pathway; positive regulation of smooth muscle cell migration; response to estradiol stimulus; response to insulin stimulus; platelet degranulation; positive regulation of MAPKKK cascade; positive regulation of glomerular filtration; positive regulation of cell proliferation; response to wounding; hemopoiesis; DNA replication; negative regulation of cell migration; epidermal growth factor receptor signaling pathway; phosphoinositide-mediated signaling; negative regulation of phosphatidylinositol biosynthetic process; positive regulation of phosphoinositide 3-kinase activity; peptidyl-serine phosphorylation; response to hypoxia; innate immune response; blood coagulation; positive regulation of DNA replication; positive regulation of cell migration Disease: Meningioma, Familial, Susceptibility To; Dermatofibrosarcoma Protuberans; Basal Ganglia Calcification, Idiopathic, 5; Basal Ganglia Calcification, Idiopathic, 1 |
| NCBI Summary: | The protein encoded by this gene is a member of the platelet-derived growth factor family. The four members of this family are mitogenic factors for cells of mesenchymal origin and are characterized by a motif of eight cysteines. This gene product can exist either as a homodimer (PDGF-BB) or as a heterodimer with the platelet-derived growth factor alpha polypeptide (PDGF-AB), where the dimers are connected by disulfide bonds. Mutations in this gene are associated with meningioma. Reciprocal translocations between chromosomes 22 and 17, at sites where this gene and that for collagen type 1, alpha 1 are located, are associated with a particular type of skin tumor called dermatofibrosarcoma protuberans resulting from unregulated expression of growth factor. Two alternatively spliced transcript variants encoding different isoforms have been identified for this gene. [provided by RefSeq, Feb 2013] |
| UniProt Code: | P01127 |
| NCBI GenInfo Identifier: | 129724 |
| NCBI Gene ID: | 5155 |
| NCBI Accession: | P01127. 1 |
| UniProt Secondary Accession: | P01127,P78431, Q15354, Q6FHE7, Q9UF23, G3XAG8, |
| UniProt Related Accession: | P01127 |
| Molecular Weight: | 241 |
| NCBI Full Name: | Platelet-derived growth factor subunit B |
| NCBI Synonym Full Names: | platelet-derived growth factor beta polypeptide |
| NCBI Official Symbol: | PDGFB |
| NCBI Official Synonym Symbols: | SIS; SSV; IBGC5; PDGF2; c-sis; PDGF-2 |
| NCBI Protein Information: | platelet-derived growth factor subunit B; becaplermin; PDGF, B chain; PDGF subunit B; proto-oncogene c-Sis; platelet-derived growth factor 2; platelet-derived growth factor B chain; platelet-derived growth factor, beta polypeptide (oncogene SIS); platelet-derived growth factor beta polypeptide (simian sarcoma viral (v-sis) oncogene homolog) |
| UniProt Protein Name: | Platelet-derived growth factor subunit B |
| UniProt Synonym Protein Names: | PDGF-2; Platelet-derived growth factor B chain; Platelet-derived growth factor beta polypeptide; Proto-oncogene c-Sis; INN: Becaplermin |
| Protein Family: | Platelet-derived growth factor |
| UniProt Gene Name: | PDGFB |
| UniProt Entry Name: | PDGFB_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 (pg/mL) | O.D | Average | Corrected |
| 1500 | 2.484 2.528 | 2.506 | 2.417 |
| 750 | 1.718 1.758 | 1.738 | 1.649 |
| 375 | 0.973 0.969 | 0.971 | 0.882 |
| 187.5 | 0.471 0.485 | 0.478 | 0.389 |
| 93.75 | 0.277 0.257 | 0.267 | 0.178 |
| 46.88 | 0.209 0.181 | 0.195 | 0.106 |
| 23.44 | 0.14 0.146 | 0.143 | 0.054 |
| 0 | 0.087 0.091 | 0.089 | -- |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human PDGFB were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human PDGFB 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) | 75.62 | 126.06 | 688.77 | 82.35 | 117.37 | 630.27 |
| Standard deviation | 4.11 | 6.77 | 36.92 | 5.69 | 6.82 | 31.26 |
| C V (%) | 5.44 | 5.37 | 5.36 | 6.91 | 5.81 | 4.96 |
Recovery
The recovery of Human PDGFB 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) | 94-105 | 99 |
| EDTA plasma (n=5) | 93-106 | 99 |
| Cell culture media (n=5) | 91-105 | 96 |
Linearity
Samples were spiked with high concentrations of Human PDGFB 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 (%) | 96-108 | 94-107 | 95-108 |
| Average (%) | 103 | 100 | 102 | |
| 1:4 | Range (%) | 92-108 | 85-99 | 84-99 |
| Average (%) | 99 | 91 | 91 | |
| 1:8 | Range (%) | 90-104 | 82-92 | 83-95 |
| Average (%) | 97 | 87 | 88 | |
| 1:16 | Range (%) | 90-105 | 79-91 | 81-94 |
| Average (%) | 97 | 86 | 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 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 90µL 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.
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