P tyr 100

Manufactured by Cell Signaling Technology

Sourced in United States

The P-Tyr-100 is a monoclonal antibody that recognizes phosphorylated tyrosine residues. It is designed for use in various laboratory techniques, such as Western blotting, immunoprecipitation, and immunohistochemistry, to detect and analyze proteins with phosphorylated tyrosine residues.

Automatically generated - may contain errors

Lab products found in correlation

Close spelling variants of this product

Anti-phospho-Tyrosine mouse mAb clone P-Tyr-100 Monoclonal phospho-tyrosine antibody (p-Tyr-100) PTyr (P-Tyr-100) P-Tyr-100 #9411 Anti-P-Tyrosine (P-Tyr-100) PhosphoScan kit (P-Tyr-100) Phospho-Tyrosine Mouse mAb (P-Tyr-100) Phosphotyrosine clone P-Tyr-100 9411 P-Tyr-100 phosphotyrosine antibody beads Anti-phospho-tyrosine (p-Tyr-100),

17 protocols using p tyr 100

Anti-FLAG and Phospho-Specific Antibody Protocol

Check if the same lab product or an alternative is used in the 5 most similar protocols

Anti-FLAG M2 (#F1804) and Anti-FLAG M2-HRP (#A8592) were purchased from Sigma-Aldrich and anti-Src antibody (#MA5-15120) was from Thermo Scientific. p-Src (Y416) (#2113), p-Abl (Y412) (#2865), p-Tyr-100 (#9411), p-Tyr-1000 (#8954), anti-GAPDH (#14C10), Myc Tag (mouse #2276 and rabbit #71D10), p-AKT (T308) (#13038), p-AKT (S473) (#4060), p70 S6 kinase (#9202), and phospho-p70 S6K (T389) (#9234) primary antibodies were purchased from Cell Signaling Technologies (CST). Secondary antibodies Goat anti-Mouse-HRP (#31430) and Goat anti-Rabbit-HRP (#31460) were from Thermo Scientific. Alexa Fluor 488 anti-mouse (#A11029) and Alexa Fluor 647 anti-rabbit (#A21245) were from Life Technologies. All primary antibodies were used at a 1:1000 dilution and secondaries at 1:5000. Protease inhibitor cocktail (#P8340), Phosphatase inhibitor cocktail 2 (#P0044) and 3 (#P5726) were purchased from Sigma-Aldrich. NeuCode labeling reagents L-Lysine:2HCl (3,3,4,4,5,5,6,6-D8, 98%) (#DLM-2641-0) and L-Lysine:2HCl (13C6, 99%, 15N2, 99%) (#CNLM-291-H-0.25) were from Cambridge Isotopes. Rapamycin was from Cayman Chemical and M-PER extraction reagent (#78501) was from Thermo Scientific. All restriction enzymes and DNA polymerases were purchased from NEB (Ipswich, MA). Oligonucleotides and gBlocks Gene Fragments were purchased from IDT and all constructs were verified by DNA sequencing (Quintara Biosciences).

Diaz J.E., Morgan C.W., Minogue C.E., Hebert A.S., Coon J.J, & Wells J.A. (2017). A Split-Abl Kinase for Direct Activation in Cells. Cell chemical biology, 24(10), 1250-1258.e4.

+ Open protocol

+ Expand

Phosphorylation Detection in Cell Biology

Cited 3 times

Check if the same lab product or an alternative is used in the 5 most similar protocols

For KiPIK screening, immunoblotting and immunofluorescence microscopy, rabbit polyclonal antibodies to H3T3ph (B8634)^{47 (link)}, INCENP (P240, Cell Signaling Technology #2807), INCENP-S446ph (Jan-Michael Peters, IMP, Vienna)^{57 (link)}, INCENP-TSSph (Michael Lampson, University of Pennsylvania)^{82 (link)}, BCL9L S915ph (Cell Signaling Technology #13325), Neurogranin S36ph (Merck-Millipore, ABN426) and γ-tubulin (AK-15, Sigma, T3320); rabbit monoclonal antibodies to the S-pT-P motif (D73F6; Cell Signaling Technology #5243), Cyclin B1 (D5C10; Cell Signaling Technology #12231) and Vinculin (E1E9V; Cell Signaling Technology #13901); mouse monoclonal antibodies to H3S28ph (CMA315)^{83 (link)} and phospho-tyrosine (P-Tyr-100; Cell Signaling Technology #9411); and sheep polyclonal antibodies to Aurora B (SAB.1, Stephen Taylor, University of Manchester)^{84 (link)} and BCL9L (R&D Systems, AF4967) were used. Antibodies used for siRNA screening were mouse monoclonal anti-H3T3ph (16B2)^{85 (link)} and rabbit polyclonal anti-H2BS6ph^{86 (link)}. Secondary antibodies were: donkey anti-sheep IgG-HRP (ThermoFisher, A16041), goat anti-rabbit IgG-HRP (Cell Signaling Technology #7074), horse anti-mouse IgG-HRP (Cell Signaling Technology #7076), donkey anti-mouse Alexa Fluor 488, anti-rabbit Alexa Fluor 594 and anti-sheep Alexa 488 Fluor (ThermoFisher, A-21202, A-21207, and A-11015).

Watson N.A., Cartwright T.N., Lawless C., Cámara-Donoso M., Sen O., Sako K., Hirota T., Kimura H, & Higgins J.M. (2020). Kinase inhibition profiles as a tool to identify kinases for specific phosphorylation sites. Nature Communications, 11, 1684.

+ Open protocol

+ Expand

Collagen I Substrate-Mediated AXL Signaling

Check if the same lab product or an alternative is used in the 5 most similar protocols

Coverslips were coated with 100 µg/mL collagen I in 20 mM acetic acid, then seeded with cells in the presence or absence of warfarin. Cells were fixed 24 hrs later with 4% paraformaldehyde in PHEM buffer. Fixed cells were permeabilized with 0.1% Triton X-100 and 1% BSA in PBS, then blocked 30 min using Odyssey blocking buffer (OBB; Li-Cor Corp.). The primary antibody (R&D Systems: AXL mAb, MAB154, 5 µg/mL) was incubated for 1 hour at room temperature in OBB. After washing with 1% BSA in PBS, cells were incubated with phalloidin and secondary antibodies for 1 hour and finally washed with 1% BSA in PBS.
For bead-based stimulation, Gas6 was coupled to polystyrene beads following identical methods to the capture antibodies. Coupled beads were washed six times to remove uncoupled protein. Cells were starved for 4 hr, and then beads were added and allowed to settle for the indicated amount of time. 4% PFA in PHEM buffer with phosphatase inhibitor (Boston Bioproducts) was used to fix the cells for 10 min. Immunofluorescence was performed by standard methods, with 1 µg/mL phosphotyrosine antibody (P-Tyr-100, Cell Signaling Technology).
Imaging was performed using a CARVII spinning disk confocal microscope with a 40× objective. Stacks were imaged every 1 µm, and then processed by maximum projection.

Meyer A.S., Zweemer A.J, & Lauffenburger D.A. (2015). The AXL Receptor is a Sensor of Ligand Spatial Heterogeneity. Cell systems, 1(1), 25-36.

+ Open protocol

+ Expand

Insulin-induced EmIR1 tyrosine phosphorylation

Check if the same lab product or an alternative is used in the 5 most similar protocols

Intact in vitro cultivated metacestode vesicles (H95) were manually picked, transferred into Falcon tubes and incubated in (D)MEM (0.2% FCS) in the presence or absence of 100 nM insulin (saturating concentration to ensure maximal activation [25 (link)-30 (link)]). After 10 minutes incubation, medium was removed and the metacestode vesicles were mechanically disrupted and pelleted by centrifugation (one minute, 1,300 rpm, 4°C). The pellet was resuspended in 1 ml lysis buffer (see above), agitated for one hour at 4°C and insoluble material was removed by centrifugation (15 minutes, 1,300 rpm, 4°C). Immunoprecipitation of EmIR1 using the anti-EmIR1 antiserum was carried out as described above and precipitated proteins were analysed by Western blotting with the anti-EmIR1 antiserum. Tyrosine phosphorylation of the EmIR1 β-subunit was carried out using an anti-phospho-tyrosine antibody (P-Tyr 100, Cell Signalling).

Hemer S., Konrad C., Spiliotis M., Koziol U., Schaack D., Förster S., Gelmedin V., Stadelmann B., Dandekar T., Hemphill A, & Brehm K. (2014). Host insulin stimulates Echinococcus multilocularis insulin signalling pathways and larval development. BMC Biology, 12, 5.

+ Open protocol

+ Expand

FHL1 Phosphorylation Signaling Pathways

Check if the same lab product or an alternative is used in the 5 most similar protocols

FHL1 (rabbit) antibody was purchased from ProteinTech. BCLAF1 (rabbit) and kindlin-2 (mouse) antibodies were from EMD Millipore. kindlin-2 (rabbit), Flag (mouse), GFP (mouse), and HA (mouse) antibodies as well as anti-flag M2 beads were purchased from Sigma-Aldrich. Src (mouse), β-actin (mouse), YY1 (mouse), and β-tubulin (mouse) were obtained from Santa Cruz Biotechnology, Inc. Anti–p-Src Y416 and –p-Tyr–100 specifically recognizing phosphorylated tyrosine were purchased from Cell Signaling Technology. Antibodies specifically recognizing Y149- and Y272-phosphorylated FHL1 were produced by immunizing rabbits with phosphorylated peptides FFPKGEDFY^PCVTC and CHQEQVY^PCPDCAKK, respectively (Kang Wei Shi Ji). Secondary antibodies conjugated with Alexa Fluor 488, 568, or 633 for immunofluorescence were purchased from Invitrogen. Src family kinase inhibitor PP2 was purchased from Sigma-Aldrich. λ-Phosphatase (P0753S) was obtained from New England Biolabs, Inc.

Wang X., Wei X., Yuan Y., Sun Q., Zhan J., Zhang J., Tang Y., Li F., Ding L., Ye Q, & Zhang H. (2018). Src-mediated phosphorylation converts FHL1 from tumor suppressor to tumor promoter. The Journal of Cell Biology, 217(4), 1335-1351.

+ Open protocol

+ Expand

Phosphotyrosine Peptide Enrichment and Quantification

Check if the same lab product or an alternative is used in the 5 most similar protocols

Cells were lysed in denaturing buffer then proteins were reduced, alkylated and digested with trypsin. The tryptic peptides were then desalted and lyophilized, with enrichment for phosphotyrosine-containing peptides using immunoprecipitation with immobilized anti-phosphotyrosine antibody p-Tyr-100 (Cell Signaling Technology). A nanoflow ultra high performance liquid chromatograph (RSLC, Dionex, Sunnyvale, CA) coupled to an electrospray ion trap mass spectrometer (LTQ-Orbitrap, Thermo, San Jose, CA) was used for tandem mass spectrometry. After Sequest and Mascot searches, the results were summarized in Scaffold 3.0. The relative quantification of tyrosine-phosphorylated peptides was calculated using MaxQuant (ver 1.2.2.5). Multi Experiment Viewer was used for generating a heatmap of the relative pY intensities (version 4.8.1) and performing Pearson correlation analysis between NRAS- and BRAF-mutant cell line cohorts.

Fedorenko I.V., Fang B., Koomen J.M., Gibney G.T, & Smalley K.S. (2014). Amuvatinib has cytotoxic effects against NRAS- but not BRAF-mutant melanoma. Melanoma research, 24(5), 448-453.

+ Open protocol

+ Expand

Syk Phosphorylation Analysis by Western Blot

Check if the same lab product or an alternative is used in the 5 most similar protocols

Phosphorylation of Syk in the 25 μg cytoplasmic fraction in NP1 lysis buffer (Cell Signaling Technology) was analyzed by Western blotting using monoclonal phospho-Syk (Tyr525/526) (C87C1), or polyclonal phospho-Zap-70 (Tyr319)/Syk (Tyr352) Ab and Syk Ab, all from Cell Signaling Technology. In some experiments, the whole cell lysate was immunoprecipitated with phospho-Tyr mouse mAb (P-Tyr-100, Cell Signaling Technology) according to the manufacturer´s instructions and the immunoprecipitate was analyzed by Western blotting using Syk Ab.

Aouar B., Kovarova D., Letard S., Font-Haro A., Florentin J., Weber J., Durantel D., Chaperot L., Plumas J., Trejbalova K., Hejnar J., Nunès J.A., Olive D., Dubreuil P., Hirsch I, & Stranska R. (2016). Dual Role of the Tyrosine Kinase Syk in Regulation of Toll-Like Receptor Signaling in Plasmacytoid Dendritic Cells. PLoS ONE, 11(6), e0156063.

+ Open protocol

+ Expand

Enrichment and Analysis of Phosphopeptides

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

Samples for peptide analysis were prepared as previously described^{1 (link)}. Samples for phosphopeptide analysis were prepared as previously described^{37 (link)} with the following modifications. Following tandem mass tag (TMT) labeling (1:4; peptide:TMT label, Pierce), phospho-tyrosine (pY) peptides were enriched from the total phosphopeptide pool using pY-specific antibodies as described by the manufacturer (Cell Signaling, P-Tyr-100). The pY enriched peptide data was collected as a single 3 hour run on the mass spectrometer. The flow through from the pY-enrichment was used to collect the phospho-serine/threonine data as previously described^{37 (link)}.

Nabet B., Roberts J.M., Buckley D.L., Paulk J., Dastjerdi S., Yang A., Leggett A.L., Erb M.A., Lawlor M.A., Souza A., Scott T.G., Vittori S., Perry J.A., Qi J., Winter G.E., Wong K.K., Gray N.S, & Bradner J.E. (2018). The dTAG system for immediate and target-specific protein degradation. Nature chemical biology, 14(5), 431-441.

+ Open protocol

+ Expand

Immunoprecipitation of Nuclear Proteins

Check if the same lab product or an alternative is used in the 5 most similar protocols

Immunoprecipitation was performed as described previously, with some modification [47 (link), 48 ]. Briefly, protein was extracted from nuclear pellets with IP buffer and diluted to 50 mmol/L NaCl, then pre-cleared with IgG and protein A/G PLUS agarose beads (Santa Cruz). Cleared lysates were incubated with specific antibody or IgG overnight with agitation at 4°C. Immune complexes were precipitated with protein A/G PLUS beads, washed, and subjected to SDS-PAGE and immunoblotting analysis. Specific antibodies used for antigen capture in IP studies were Src (B-12), p300 (C20) (Santa Cruz), and p-TYR-100 (Cell Signaling).

Paladino D., Yue P., Furuya H., Acoba J., Rosser C.J, & Turkson J. (2015). A novel nuclear Src and p300 signaling axis controls migratory and invasive behavior in pancreatic cancer. Oncotarget, 7(6), 7253-7267.

+ Open protocol

+ Expand

Culturing Murine Fibroblasts and Melanoma Cells

Cited 1 time

Check if the same lab product or an alternative is used in the 5 most similar protocols

Murine fibroblasts (NR6WT) expressing human epidermal growth factor receptor and modified as previously described^{4 (link)}, were grown in alpha-MEM media supplemented with 1x non-essential amino acids, 1x sodium pyruvate, 1x L-glutamine, 1x pen/strep antibiotics, and 7.5% fetal bovine serum. Parental Melanoma cell line WM1158 and ACTN4 knockdown WM1158 (WM1158 ACTN4 KD)^{20 (link)} cells were grown in DMEM (1 gL⁻¹ glucose):L15 3:1 medium with 10% fetal bovine serum and 1x pen/strip antibiotics. Transfection reagent xFect was purchased from Clontech Life Technologies (Grand Island, NY). Monoclonal antibody against phosphorylated tyrosine (P-Tyr-100) and polyclonal GAPDH antibody were purchased from Cell Signaling Technology (Beverly, MA). Polyclonal GFP antibody was purchased from Abcam (Cambridge MA). Purified bovine muscle actin and polyclonal Actin antibody were purchased from Sigma Aldrich (St. Louis, MO).

Shao H., Wingert B., Weins A., Pollak M.R., Camacho C, & Wells A. (2019). Focal segmental glomerulosclerosis ACTN4 mutants binding to actin: regulation by phosphomimetic mutations. Scientific Reports, 9, 15517.

+ Open protocol

+ Expand

About PubCompare

Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.

We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.

However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.

Ready to get started?

Revolutionizing how scientists
search and build protocols!