Revision 6

#9411

Store at -20C

Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Orders:

877-616-CELL (2355)

[email protected]

Support:

877-678-TECH (8324)

Web:

[email protected]

cellsignal.com

3 Trask Lane | Danvers | Massachusetts | 01923 | USA

For Research Use Only. Not for Use in Diagnostic Procedures.

Applications:
W, IP, IHC-P, IF-F, IF-IC, FC-FP, E-P

Reactivity:
All

Sensitivity:
Endogenous

Source/Isotype:
Mouse IgG1

Product Usage Information

The phosphorylated form of the peptide can be detected with Phospho-Tyrosine mAb (P-Tyr-100) #9411. Sample kinase protocol is attached.

Application	Dilution
Western Blotting	1:2000
Immunoprecipitation	1:100
Immunohistochemistry (Paraffin)	1:2400 - 1:9600
Immunofluorescence (Frozen)	1:1600 - 1:3200
Immunofluorescence (Immunocytochemistry)	1:1600 - 1:3200
Flow Cytometry (Fixed/Permeabilized)	1:1600 - 1:6400
Peptide ELISA (DELFIA)	1:4000

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

For a carrier free (BSA and azide free) version of this product see product #80379.

Specificity/Sensitivity

Phospho-Tyrosine Mouse mAb (P-Tyr-100) is a high affinity antibody. ELISAs against a wide variety of phosphopeptides indicate that P-Tyr-100 binds phospho-Tyr in a manner largely independent of the surrounding amino acid sequence. 2D gel Western blot analysis of pervanadate-treated cell extracts also shows that P-Tyr-100 interacts with a broad range of tyrosine-phosphorylated proteins. P-Tyr-100 does not cross-react with peptides containing phospho-Ser or phospho-Thr.

Source / Purification

Monoclonal antibody is produced by immunizing animals with phospho-tyrosine containing peptides .

Background

Tyrosine phosphorylation plays a key role in cellular signaling (1). Research studies have shown that in cancer, unregulated tyrosine kinase activity can drive malignancy and tumor formation by generating inappropriate proliferation and survival signals (2). Antibodies specific for phospho-tyrosine (3,4) have been invaluable reagents in these studies. The phospho-tyrosine monoclonal antibodies developed by Cell Signaling Technology are exceptionally sensitive tools for studying tyrosine phosphorylation and monitoring tyrosine kinase activity in high throughput drug discovery.

Background References

Species Reactivity

Species reactivity is determined by testing in at least one approved application (e.g., western blot).

Western Blot Buffer

IMPORTANT: For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.

Applications Key

W: Western Blotting IP: Immunoprecipitation IHC-P: Immunohistochemistry (Paraffin) IF-F: Immunofluorescence (Frozen) IF-IC: Immunofluorescence (Immunocytochemistry) FC-FP: Flow Cytometry (Fixed/Permeabilized) E-P: Peptide ELISA (DELFIA)

Cross-Reactivity Key

All: All Species Expected

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.

All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

限制使用

除非 CST 的合法授书代表以书面形式书行明确同意，否书以下条款适用于 CST、其关书方或分书商提供的书品。任何书充本条款或与本条款不同的客书条款和条件，除非书 CST 的合法授书代表以书面形式书独接受，否书均被拒书，并且无效。

专品专有“专供研究使用”的专专或专似的专专声明，且未专得美国食品和专品管理局或其他外国或国内专管机专专专任何用途的批准、准专或专可。客专不得将任何专品用于任何专断或治专目的，或以任何不符合专专声明的方式使用专品。CST 专售或专可的专品提供专作专最专用专的客专，且专用于研专用途。将专品用于专断、专防或治专目的，或专专售（专独或作专专成）或其他商专目的而专专专品，均需要 CST 的专独专可。客专：(a) 不得专独或与其他材料专合向任何第三方出售、专可、出借、捐专或以其他方式专专或提供任何专品，或使用专品制造任何商专专品，(b) 不得复制、修改、逆向工程、反专专、反专专专品或以其他方式专专专专专品的基专专专或技专，或使用专品开专任何与 CST 的专品或服专专争的专品或服专， (c) 不得更改或专除专品上的任何商专、商品名称、徽专、专利或版专声明或专专，(d) 只能根据 CST 的专品专售条款和任何适用文档使用专品， (e) 专遵守客专与专品一起使用的任何第三方专品或服专的任何专可、服专条款或专似专专

Orders: 877-616-CELL (2355) • [email protected] • Support: 877-678-TECH (8324) • [email protected] • Web: cellsignal.com

For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 6

#9411

Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Western blot analysis of extracts from Jurkat cells treated with 1 mM pervanadate for 30 minutes prior to lysis, using P-Tyr-100 Phospho-Tyrosine Mouse mAb. Proteins were separated by 2-D electrophoresis prior to blotting.

Western Blotting Image 1: Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Immunohistochemical analysis of paraffin-embedded human breast carcinoma using Phospho-Tyrosine Mouse mAb (P-Tyr-100).

Immunohistochemistry Image 1: Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Immunohistochemical analysis of paraffin-embedded human lung carcinoma using Phospho-Tyrosine Mouse mAb (P-Tyr-100).

Immunohistochemistry Image 2: Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Orders: 877-616-CELL (2355) • [email protected] • Support: 877-678-TECH (8324) • [email protected] • Web: cellsignal.com

For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 6

#9411

Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Immunohistochemical analysis of paraffin-embedded NCI-H1650 xenograft untreated (left) or lambda-phosphatase-treated (right), using Phospho-Tyrosine Mouse mAb (P-Tyr-100).

Immunohistochemistry Image 3: Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Immunohistochemical analysis of paraffin-embedded human B-cell non-Hodgkin lymphoma, untreated (left) or lambda phosphatase treated (right), using Phospho-Tyrosine Mouse mAb (P-Tyr-100).

Immunohistochemistry Image 4: Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Immunohistochemical analysis of paraffin-embedded human squamous cell lung carcinoma using Phospho-Tyrosine Mouse mAb (P-Tyr-100).

Immunohistochemistry Image 5: Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Orders: 877-616-CELL (2355) • [email protected] • Support: 877-678-TECH (8324) • [email protected] • Web: cellsignal.com

For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 6

#9411

Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Immunohistochemical analysis using Phospho-Tyrosine Mouse mAb (P-Tyr-100) on SignalSlide™ Phospho-EGF Receptor IHC Controls #8102 (paraffin-embedded KYSE450 cell pellets, untreated (left) or EGF-treated (right)).

Immunohistochemistry Image 6: Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Immunohistochemical analysis of paraffin-embedded human soft tissue squamous cell carcinoma using Phospho-Tyrosine Mouse mAb (P-Tyr-100).

Immunohistochemistry Image 7: Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Immunohistochemical analysis of paraffin-embedded normal human kidney using Phospho-Tyrosine Mouse mAb (P-Tyr-100).

Immunohistochemistry Image 8: Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Orders: 877-616-CELL (2355) • [email protected] • Support: 877-678-TECH (8324) • [email protected] • Web: cellsignal.com

For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 6

#9411

Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Immunohistochemical analysis of paraffin-embedded normal human lung using Phospho-Tyrosine Mouse mAb (P-Tyr-100).

Immunohistochemistry Image 9: Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Immunohistochemical analysis of paraffin-embedded NCI-H1650 xenograft using Phospho-Tyrosine Mouse mAb (P-Tyr-100).

Immunohistochemistry Image 10: Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Immunofluorescent analysis of Swiss NIH/3T3 cells, serum-starved and stimulated with lysophosphatidic acid (LPA) (10 µM for 10 minutes) and fixed with PFA, using Phospho-Tyrosine Mouse mAb (P-Tyr-100) (red) and phalloidin for F-actin (green) LPA causes heavy tyrosine phosphorylation of proteins in focal adhesions, present at the tips of actin stress fibers. (Provided by Dr. Harry Mellor, University of Bristol.)

Immunofluorescence Image 1: Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Orders: 877-616-CELL (2355) • [email protected] • Support: 877-678-TECH (8324) • [email protected] • Web: cellsignal.com

For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 6

#9411

Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Confocal immunofluorescence analysis of HeLa cells either stimulated with 20% serum (left) or untreated (right), using Phospho-Tyrosine Mouse mAb (P-Tyr-100) (red). Actin filaments have been labeled with fluorescein phalloidin. Blue pseudocolor = DRAQ5^® (fluorescent DNA dye).

Immunofluorescence Image 2: Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Flow cytometric analysis of K562 cells, untreated (green) or Gleevec^®- treated (blue), using Phospho-Tyrosine Mouse mAb (P-Tyr-100) compared to a nonspecific negative control antibody (red).

Flow Cytometry Image 1: Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Phospho-Tyrosine Mouse mAb (P-Tyr-100) (HRP Conjugate) ELISA Assay: Signal-to-noise ratio of phospho- versus nonphospho-peptides. (Y* denotes phosphorylated tyrosine.)

E Image 1: Phospho-Tyrosine Mouse mAb (P-Tyr-100)

Orders: 877-616-CELL (2355) • [email protected] • Support: 877-678-TECH (8324) • [email protected] • Web: cellsignal.com

For Research Use Only. Not for Use in Diagnostic Procedures.