Human TNK2 (Tyrosine Kinase, Non Receptor 2) ELISA Kit (HUES02892)

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HUES02892
€539

Description

ELISA Kit Technical ManualMSDS

Human TNK2 (Tyrosine Kinase, Non Receptor 2) ELISA Kit

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 TNK2 . 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 TNK2 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 TNK2, 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 TNK2. The concentration of Human TNK2 in samples can be calculated by comparing the OD of the samples to the standard curve.

Assay typeSandwich
Format96T
Assay time4.5h
ReactivityHuman
Detection MethodColormetric
Detection Range0.31-20 ng/mL
Sensitivity0.19 ng/mL
Sample Volume Required Per Well100uL
Sample TypeSerum, plasma and other biological fluids

Specificity

This kit recognizes Human TNK2 in samples. No significant cross-reactivity or interference between Human TNK2 and analogues was observed.

Typical Data

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
202.283
2.287
2.2852.211
101.567
1.605
1.5861.512
50.828
0.804
0.8160.742
2.50.436
0.444
0.440.366
1.250.253
0.247
0.250.176
0.630.174
0.156
0.1650.091
0.310.117
0.125
0.1210.047
00.073
0.075
0.074--

Precision

Intra-assay Precision (Precision within an assay): 3 samples with low, mid range and high level Human TNK2 were tested 20 times on one plate, respectively.

Inter-assay Precision (Precision between assays): 3 samples with low, mid range and high level Human TNK2 were tested on 3 different plates, 20 replicates in each plate.

Intra-assay Precision Inter-assay Precision
Sample123123
n202020202020
Mean (ng/mL)1.081.789.811.131.848.91
Standard deviation0.070.100.470.060.100.29
C V (%)6.485.624.795.315.433.25

Recovery

The recovery of Human TNK2 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)90-10495
EDTA plasma (n=5)91-10398
Cell culture media (n=5)90-10297

Linearity

Samples were spiked with high concentrations of Human TNK2 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)
1:2Range (%)93-11095-11097-113
Average (%)101100105
1:4Range (%)94-10581-9288-99
Average (%)998794
1:8Range (%)91-10486-9982-94
Average (%)989288
1:16Range (%)87-9984-9587-99
Average (%)938993

Kit Components & Storage

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 X12 strips-20'C, 6 months
Reference Standard2 vials
Concentrated Biotinylated Detection Ab (100X)1 vial, 120 uL
Concentrated HRP Conjugate (100X)1 vial, 120 uL-20'C(shading light), 6 months
Reference Standard & Sample Diluent1 vial, 20 mL4'C, 6 months
Biotinylated Detection Ab Diluent1 vial, 14 mL
HRP Conjugate Diluent1 vial, 14 mL
Concentrated Wash Buffer (25X)1 vial, 30 mL
Substrate Reagent1 vial, 10 mL4'C(shading light)
Stop Solution1 vial, 10 mL4'C
Plate Sealer5 pieces
Product Description1 copy
Certificate of Analysis1 copy

Human TNK2 (Tyrosine Kinase, Non Receptor 2) ELISA Kit (HUES02892) Assay procedure

    1. 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.
    2. Aliquot 100µl of standard solutions into the standard wells.
    3. Add 100µl of Sample / Standard dilution buffer into the control (zero) well.
    4. Add 100µl of properly diluted sample (serum, plasma, tissue homogenates and otherbiological fluids) into test sample wells.
    5. Cover the plate with the sealer provided in the kit and incubate for 90 min at 37°C.
    6. 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.
    7. 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.
    8. Add 100µL of HRP Conjugate working solution to each well. Cover with a plate seal andincubate for 30 min at 37°C.
    9. Aspirate or decant the solution from each well. Repeat the wash process for five times asconducted in step 7.
    10. 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.
    11. Add 50 µL of Stop Solution to each well. Note: Adding the stop solution should be done inthe same order as the substrate solution.
    12. Determine the optical density (OD value) of each well immediately with a microplate readerset at 450 nm.

Human TNK2 (Tyrosine Kinase, Non Receptor 2) ELISA Kit (HUES02892) Protein Information

UniProt Protein Function:Ack: a tyrosine kinase that binds Cdc42 in its GTP-bound form and inhibits both the intrinsic and GTPase-activating protein (GAP)-stimulated GTPase activity of Cdc42. This binding is mediated by a unique sequence of 47 amino acids C-terminal to an SH3 domain. May be involved in a regulatory mechanism that sustains the GTP-bound active form of Cdc42 and which is directly linked to a tyrosine phosphorylation signal transduction pathway. Amplification in primary tumors correlates with metastatic potential. Two differentially spliced isoforms have been described.
UniProt Protein Details:

Protein type:Kinase, protein; EC 2.7.11.1; EC 2.7.10.2; Protein kinase, tyrosine (non-receptor); Protein kinase, TK; TK group; Ack family

Chromosomal Location of Human Ortholog: 3q29

Cellular Component: clathrin-coated vesicle; coated pit; cytoplasm; endosome; extrinsic to internal side of plasma membrane; membrane; nucleus; plasma membrane

Molecular Function:epidermal growth factor receptor binding; GTPase inhibitor activity; identical protein binding; protein binding; protein serine/threonine/tyrosine kinase activity; protein-tyrosine kinase activity; WW domain binding

Biological Process: cell differentiation; cell migration; cell surface receptor linked signal transduction; innate immune response; phosphorylation; positive regulation of peptidyl-tyrosine phosphorylation; regulation of cell proliferation; small GTPase mediated signal transduction; transmembrane receptor protein tyrosine kinase signaling pathway

NCBI Summary:This gene encodes a tyrosine kinase that binds Cdc42Hs in its GTP-bound form and inhibits both the intrinsic and GTPase-activating protein (GAP)-stimulated GTPase activity of Cdc42Hs. This binding is mediated by a unique sequence of 47 amino acids C-terminal to an SH3 domain. The protein may be involved in a regulatory mechanism that sustains the GTP-bound active form of Cdc42Hs and which is directly linked to a tyrosine phosphorylation signal transduction pathway. Several alternatively spliced transcript variants have been identified from this gene, but the full-length nature of only two transcript variants has been determined. [provided by RefSeq, Jul 2008]
UniProt Code:Q07912
NCBI GenInfo Identifier:229462980
NCBI Gene ID:10188
NCBI Accession:Q07912.3
UniProt Secondary Accession:Q07912,Q6ZMQ0, Q8N6U7, Q96H59,
UniProt Related Accession:Q07912
Molecular Weight:119,349 Da
NCBI Full Name:Activated CDC42 kinase 1
NCBI Synonym Full Names:tyrosine kinase non receptor 2
NCBI Official Symbol:TNK2  
NCBI Official Synonym Symbols:ACK; ACK1; ACK-1; p21cdc42Hs  
NCBI Protein Information:activated CDC42 kinase 1
UniProt Protein Name:Activated CDC42 kinase 1
UniProt Synonym Protein Names:Tyrosine kinase non-receptor protein 2
Protein Family:Activated CDC42 kinase
UniProt Gene Name:TNK2  
UniProt Entry Name:ACK1_HUMAN
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Additional Information

Product type:
ELISA
Reactivity:
Human
ELISA Type:
Sandwich
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