Human FAK (Focal Adhesion Kinase) ELISA Kit (HUES02747)
- SKU:
- HUES02747
- Product Type:
- ELISA Kit
- Size:
- 96 Assays
- Uniprot:
- Q05397
- Sensitivity:
- 0.09ng/mL
- Range:
- 0.16-10ng/mL
- ELISA Type:
- Sandwich
- Synonyms:
- PTK2
- Reactivity:
- Human
- Sample Type:
- Serum, plasma and other biological fluids
- Research Area:
- Cardiovascular
Description
Human FAK (Focal Adhesion Kinase) ELISA Kit
The Human FAK (Focal Adhesion Kinase) ELISA Kit is a powerful tool for the quantitative measurement of FAK levels in human samples such as serum, plasma, and cell culture supernatants. With its high sensitivity and specificity, this kit provides accurate and reliable results for a variety of research applications.FAK is a key protein involved in cell adhesion and migration, playing a critical role in processes such as cell growth, differentiation, and survival. Dysregulation of FAK has been linked to various diseases including cancer, cardiovascular diseases, and inflammatory disorders, highlighting its importance as a potential therapeutic target.
By using the Human FAK ELISA Kit, researchers can gain valuable insights into the role of FAK in disease pathology and explore innovative treatment strategies. This kit is user-friendly, cost-effective, and suitable for use in both academic and clinical settings, making it a valuable asset for any laboratory studying FAK signaling pathways.
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 FAK in samples. No significant cross-reactivity or interference between Human FAK 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 FAK. 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 FAK 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 FAK, 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 FAK. The concentration of Human FAK in samples can be calculated by comparing the OD of the samples to the standard curve.
UniProt Protein Function: | FAK: a tyrosine kinase of the FAK family required for cell migration and contact-dependent survival signaling. Activated by tyrosine-phosphorylation in response to either integrin clustering induced by cell adhesion or antibody cross-linking, or via G-protein coupled receptor (GPCR) occupancy by ligands such as bombesin or lysophosphatidic acid, or via LDL receptor occupancy. Downstream of integrins and Src, upstream of Ras/MAPK. Localizes to focal adhesions that form between cells growing in the presence of extracellular matrix constituents. Interacts with CAS family members and with GIT1, SORBS1 and BCAR3. Interacts with Shb. Required for full Ras transformation of fibroblasts. Increased expression in breast and other cancers, related to chromosome 8q amplification. Overexpression and activation associated with increased migration, invasion and progression of ovarian cancer, and with progression in hepatocellular carcinoma, thyroid cancer, and acute myelogenous leukemia. siRNA increases chemosensitivity of pancreatic adenocarcinoma xenografts. Inhibitor: ISI15421 (antisense). Four splice-variant isoforms have been observed. |
UniProt Protein Details: | Protein type:Kinase, protein; Protein kinase, tyrosine (non-receptor); EC 2. 7. 10. 2; Protein kinase, TK; TK group; Fak family Chromosomal Location of Human Ortholog: 8q24. 3 Cellular Component: extrinsic to internal side of plasma membrane; cytoskeleton; focal adhesion; lamellipodium; apical plasma membrane; cytoplasm; stress fiber; plasma membrane; microtubule organizing center; cell cortex; cytosol; nucleus Molecular Function:JUN kinase binding; signal transducer activity; protein binding; protein-tyrosine kinase activity; non-membrane spanning protein tyrosine kinase activity; SH2 domain binding; actin binding; protein kinase binding; ATP binding; protein kinase activity; receptor binding Biological Process: heart morphogenesis; axon guidance; extracellular matrix organization and biogenesis; peptidyl-tyrosine phosphorylation; establishment of nucleus localization; apoptosis; protein amino acid autophosphorylation; neuron migration; cell motility involved in cell locomotion; negative regulation of synaptogenesis; regulation of cell shape; regulation of cell adhesion mediated by integrin; transforming growth factor beta receptor signaling pathway; positive regulation of cell proliferation; ephrin receptor signaling pathway; negative regulation of axonogenesis; angiogenesis; vasculogenesis; placenta development; cell structure disassembly during apoptosis; integrin-mediated signaling pathway; epidermal growth factor receptor signaling pathway; platelet activation; central nervous system neuron axonogenesis; regulation of osteoblast differentiation; positive regulation of phosphoinositide 3-kinase activity; signal complex assembly; cytoskeleton organization and biogenesis; microtubule cytoskeleton organization and biogenesis; negative regulation of organ growth; regulation of cell proliferation; positive regulation of phosphoinositide 3-kinase cascade; positive regulation of protein kinase B signaling cascade; embryonic development; establishment of cell polarity; positive regulation of protein kinase activity; regulation of focal adhesion formation; endothelial cell migration; innate immune response; positive regulation of protein amino acid phosphorylation; negative regulation of cell-cell adhesion; blood coagulation; vascular endothelial growth factor receptor signaling pathway; regulation of cytoskeleton organization and biogenesis; negative regulation of apoptosis; positive regulation of cell migration |
NCBI Summary: | This gene encodes a cytoplasmic protein tyrosine kinase which is found concentrated in the focal adhesions that form between cells growing in the presence of extracellular matrix constituents. The encoded protein is a member of the FAK subfamily of protein tyrosine kinases but lacks significant sequence similarity to kinases from other subfamilies. Activation of this gene may be an important early step in cell growth and intracellular signal transduction pathways triggered in response to certain neural peptides or to cell interactions with the extracellular matrix. Several transcript variants encoding different isoforms have been found for this gene, but the full-length natures of only three of them have been determined. [provided by RefSeq, Dec 2010] |
UniProt Code: | Q05397 |
NCBI GenInfo Identifier: | 3183518 |
NCBI Gene ID: | 5747 |
NCBI Accession: | Q05397. 2 |
UniProt Secondary Accession: | Q05397,Q14291, Q8IYN9, Q9UD85, B4E2N6, F5H4S4, J3QT16 |
UniProt Related Accession: | Q05397 |
Molecular Weight: | 1052 |
NCBI Full Name: | Focal adhesion kinase 1 |
NCBI Synonym Full Names: | protein tyrosine kinase 2 |
NCBI Official Symbol: | PTK2 |
NCBI Official Synonym Symbols: | FAK; FADK; FAK1; FRNK; PPP1R71; p125FAK; pp125FAK |
NCBI Protein Information: | focal adhesion kinase 1; FADK 1; PTK2 protein tyrosine kinase 2; FAK-related non-kinase polypeptide; focal adhesion kinase isoform FAK-Del33; focal adhesion kinase-related nonkinase; protein phosphatase 1 regulatory subunit 71; protein phosphatase 1, regulatory subunit 71 |
UniProt Protein Name: | Focal adhesion kinase 1 |
UniProt Synonym Protein Names: | Focal adhesion kinase-related nonkinase; FRNK; Protein phosphatase 1 regulatory subunit 71; PPP1R71; Protein-tyrosine kinase 2; p125FAK; pp125FAK |
Protein Family: | Focal adhesion kinase |
UniProt Gene Name: | PTK2 |
UniProt Entry Name: | FAK1_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 | 2.392 2.436 | 2.414 | 2.328 |
5 | 1.699 1.751 | 1.725 | 1.639 |
2.5 | 1.007 0.975 | 0.991 | 0.905 |
1.25 | 0.533 0.537 | 0.535 | 0.449 |
0.63 | 0.312 0.292 | 0.302 | 0.216 |
0.32 | 0.202 0.19 | 0.196 | 0.11 |
0.16 | 0.139 0.145 | 0.142 | 0.056 |
0 | 0.085 0.087 | 0.086 | -- |
Precision
Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human FAK were tested 20 times on one plate, respectively.
Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human FAK 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) | 0.49 | 1.15 | 4.03 | 0.50 | 1.14 | 4.16 |
Standard deviation | 0.03 | 0.06 | 0.16 | 0.03 | 0.06 | 0.14 |
C V (%) | 6.12 | 5.22 | 3.97 | 6.00 | 5.26 | 3.37 |
Recovery
The recovery of Human FAK 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) | 87-102 | 93 |
EDTA plasma (n=5) | 86-98 | 92 |
Cell culture media (n=5) | 85-97 | 92 |
Linearity
Samples were spiked with high concentrations of Human FAK 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 (%) | 95-109 | 83-98 | 98-110 |
Average (%) | 101 | 90 | 104 | |
1:4 | Range (%) | 90-103 | 80-93 | 81-93 |
Average (%) | 97 | 86 | 88 | |
1:8 | Range (%) | 87-99 | 80-90 | 83-97 |
Average (%) | 93 | 85 | 89 | |
1:16 | Range (%) | 89-103 | 84-97 | 85-99 |
Average (%) | 95 | 89 | 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 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.