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WB: 20 micrograms of EGF-stimulated A431 cell lysate was probed with 1µg/ml of Anti-Phosphotyrosine, clone 4G10®.
Anti-Phosphotyrosine, clone 4G10®
Recommended Replacement for: 05-321 is a recommended replacement for MAB3090| Reatividade à Espécie | Aplicações Essenciais | Hospedeiro | Formato | Tipo de Anticorpo |
|---|---|---|---|---|
| A | IC, IH, IP, WB | Mouse | Purified | Monoclonal Antibody |
Description:
Anti-Phosphotyrosine, clone 4G10®
Replaces:
05-321 is a recommended replacement for MAB3090
Background Information:
Some of the tyrosine residues can be tagged with a phosphate group (phosphorylated) by protein kinases. (In its phosphorylated state, it is referred to as phosphotyrosine.). Tyrosine phosphorylation is considered as one of the key steps in signal transduction and regulation of enzymatic activity.
The advent of anti-phosphotyrosine antibodies is one of significant events in signal transduction research. Before the availability of anti-phosphotyrosine antibodies, tyrosyl phospyhorylation of proteins and enzymes was investigated through hazardous and time-consuming radioactive experiments. Anti-phosphotyrosine antibodies are commonly used in western blots after the targeted proteins have been immunoprecipitated to measure the tyrosyl phosphorylation of the proteins. Anti-phosphotyrosine antibodies are also directly used on cell lysate to examine the overall change of tyrosine phosphorylation level in reponse to various treatments.
The advent of anti-phosphotyrosine antibodies is one of significant events in signal transduction research. Before the availability of anti-phosphotyrosine antibodies, tyrosyl phospyhorylation of proteins and enzymes was investigated through hazardous and time-consuming radioactive experiments. Anti-phosphotyrosine antibodies are commonly used in western blots after the targeted proteins have been immunoprecipitated to measure the tyrosyl phosphorylation of the proteins. Anti-phosphotyrosine antibodies are also directly used on cell lysate to examine the overall change of tyrosine phosphorylation level in reponse to various treatments.
Clone:
4G10®
Specificity:
This antibody recognizes tyrosine-phosphorylated proteins from all species.
Key Applications:
- Immunocytochemistry
- Immunohistochemistry
- Immunoprecipitation
- Western Blotting
Application Notes:
Immunoprecipitation:
2-4 μg of a previous lot immunoprecipitated quantitatively the phosphotyrosine containing proteins in the lysate of a confluent culture (10 cm dish) of cells expressing an activated tyrosine kinase. To preserve phosphotyrosine, add: 0.2 mM sodium orthovanadate to the lysis buffer.
2-4 μg of a previous lot immunoprecipitated quantitatively the phosphotyrosine containing proteins in the lysate of a confluent culture (10 cm dish) of cells expressing an activated tyrosine kinase. To preserve phosphotyrosine, add: 0.2 mM sodium orthovanadate to the lysis buffer.
Species Reactivity:
All
Species Reactivity Note:
All species
Isotype:
IgG2bκ
Immunogen:
Phosphotyramine-KLH
Modifications:
Phosphorylation
Molecular Weight:
Dependent upon the molecular weight of the tyrosine phosphorylated protein being detected.
Control:
Untreated A-431 (negative control) and EGF treated A-431 (positive control) whole cell lysates.
Included Positive Antigen Control: Catalog # 12-302, EGF-stimulated A431 cell lysate is provided as a free positive antigen control for western immunoblotting. Aliquot as desired, refreeze immediately, and store at -20°C. The lysate is stable for 6 months at -20°C. Before use, add 2.5 μL of 2-mercaptoethanol/100 μL of lysate and boil for 5 minutes to reduce the preparation. Load 20 μg of reduced lysate per lane for immunoblot analysis.
Included Positive Antigen Control: Catalog # 12-302, EGF-stimulated A431 cell lysate is provided as a free positive antigen control for western immunoblotting. Aliquot as desired, refreeze immediately, and store at -20°C. The lysate is stable for 6 months at -20°C. Before use, add 2.5 μL of 2-mercaptoethanol/100 μL of lysate and boil for 5 minutes to reduce the preparation. Load 20 μg of reduced lysate per lane for immunoblot analysis.
Presentation:
Purified mouse monoclonal IgG2bκ in buffer containing 0.02 M phosphate buffer, pH 7.6, 0.25 M NaCl, 0.1% sodium azide. Liquid at 2-8ºC.
IgG2bκ mouse monocolonal antibody produced in vitro by mouse-mouse hybridoma 4G10® (FOX-NY [NS-1 derivative] myeloma x spleen cells).
IgG2bκ mouse monocolonal antibody produced in vitro by mouse-mouse hybridoma 4G10® (FOX-NY [NS-1 derivative] myeloma x spleen cells).
Storage Conditions:
Stable for 1 year at 2-8°C from date of receipt.
NOTE: DO NOT FREEZE.
For maximum recovery of the product, centrifuge the original vial prior to removing the cap. If the product has accidentally been frozen and thawed, spin it at 13,000 x g for 10 minutes at 2-8ºC. Save the supernatant for application.
NOTE: DO NOT FREEZE.
For maximum recovery of the product, centrifuge the original vial prior to removing the cap. If the product has accidentally been frozen and thawed, spin it at 13,000 x g for 10 minutes at 2-8ºC. Save the supernatant for application.
Quality Assurance:
Routinely evaluated on EGF-treated human A431 carcinoma cells.
Western Blot Analysis:
0.5-2 μg/mL of this lot detected tyrosine-phosphorylated proteins in a modified RIPA lysate from EGF-treated human A431 carcinoma cells (Cohen, B., 1990; , Druker, B. J., 1989; Kanakura, Y., 1991).
Western Blot Analysis:
0.5-2 μg/mL of this lot detected tyrosine-phosphorylated proteins in a modified RIPA lysate from EGF-treated human A431 carcinoma cells (Cohen, B., 1990; , Druker, B. J., 1989; Kanakura, Y., 1991).
Antibody Category:
Signaling
Antibody Sub-Category:
General Post-translation Modification
Usage Statement:
Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
Trade Name:
Upstate (Millipore)
Format Code:
Pur
Concentration:
1 mg/mL
Antibody Type:
Monoclonal Antibody
Format:
Purified
Host:
Mouse
Purification Method:
Protein G-Sepharose chromatography