Anti phosphotyrosine clone 4g10 05 321

Manufactured by Merck Group

Sourced in United States

Anti-phosphotyrosine clone 4G10 (05-321) is a laboratory reagent produced by Merck Group. It functions as a monoclonal antibody that specifically recognizes and binds to phosphorylated tyrosine residues on various proteins.

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4 protocols using anti phosphotyrosine clone 4g10 05 321

Protein Purification and Immunoblotting Protocol

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Phenylmethylsulfonyl fluoride, sodium vanadate, bovine serum albumin, dithiothreitol (DTT), poly (Glu, Tyr), fluorescein isothiocyanate, holo-Transferrin, and buffer components were purchased from Sigma-Aldrich, St. Louis, MO, USA. Horseradish peroxidase linked anti-rabbit and anti-mouse secondary antibodies were from Cytiva, Marlborough, MA, USA. SuperSignal West Femto Chemiluminescent Substrate and Pierce ECL Western Blotting Substrate were purchased from Thermo Scientific, Waltham, MA, USA. Nickel-nitriloacetic acid (Ni-NTA) resin was from Qiagen, Germantown, MD, USA. The following antibodies and resin were obtained from MilliporeSigma, St. Louis, MO, USA: anti-phosphotyrosine clone 4G10 (05-321), anti-phospho-Ack1 (pY284) (09-142), anti-γ-Tubulin (T6557), horseradish peroxidase–linked anti-FLAG produced in mouse (F1804), and EZview red anti-FLAG M2 affinity resin. Anti-Mer antibody for Western blotting was from R&D Systems, Minneapolis, MN, USA (AF891). Anti-Mer antibody for immunoprecipitation was from abcam, Cambridge, UK (ab184086).

Hayashi S.Y., Craddock B.P, & Miller W.T. (2023). Phosphorylation of Ack1 by the Receptor Tyrosine Kinase Mer. Kinases and phosphatases, 1(3), 167-180.

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Protein Purification and Immunoblotting Protocol

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Hayashi S.Y., Craddock B.P, & Miller W.T. (2023). Phosphorylation of Ack1 by the Receptor Tyrosine Kinase Mer. Kinases and phosphatases, 1(3), 167-180.

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Immunofluorescence and Western Blotting Assay

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For immunofluorescence, anti-cortactin (ab-33333) was obtained from Abcam; anti–pY402-Pyk2 and anti–pY397-FAK were obtained from Invitrogen; anti-Arp2 (H-84; SC-15389) and anti-Tks5 (FISH M-300; SC-30122) were obtained from Santa Cruz Biotechnology, Inc.; and antivinculin (clone hVIN1; V9131), anti–pY421-cortactin (C0739), and anti–pY466-cortactin (C0864) were obtained from Sigma-Aldrich. Rhodamine-labeled phalloidin and Alexa Fluor–conjugated secondary antibodies were obtained from Molecular Probes. For Western blotting, anticortactin (clone 4F11; 05-180) and antiphosphotyrosine (clone 4G10; 05-321) were obtained from EMD Millipore; anti-Pyk2 (3480 and 3292) was obtained from Cell Signaling Technology; anti-FAK (clone 77; 610088) was obtained from BD; anti-GFP (clones 7.1 and 13.1; 11814460001) was obtained from Roche; and anti–β-actin (clone AC-15; A5441) was obtained from Sigma-Aldrich. Secondary antibodies (goat anti–mouse 680LT and goat anti–rabbit 800CW) were obtained from LI-COR Biosciences.

Genna A., Lapetina S., Lukic N., Twafra S., Meirson T., Sharma V.P., Condeelis J.S, & Gil-Henn H. (2018). Pyk2 and FAK differentially regulate invadopodia formation and function in breast cancer cells. The Journal of Cell Biology, 217(1), 375-395.

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Antibody Characterization for Signaling

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The following antibodies were used: anti-phospho-ERK (sc-7383) and anti-FAK (sc-557), obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA); anti-phospho-Akt (Ser473) (4060s) and anti-Akt (9272s), obtained from Cell Signaling Technology (Danvers, MA, USA); anti-phospho-tyrosine (clone 4G10) (05-321) and anti-phospho-FAK (Tyr397) (abt135), obtained from Merck Millipore (Burlington, MA, USA); anti-ERK2 (bms-52068R), obtained from Bioss (Boston, MA, USA); anti-FLAG-M2 antibody (F1804), obtained from Sigma-Aldrich; anti-Ki-67 (ab15580), obtained from Abcam (Cambridge, UK); anti-GAPDH (abc2003), obtained from AbClone (Seoul, Korea); and horseradish peroxidase-conjugated goat anti-mouse IgG (K0211589) and rabbit IgG (K0211708), obtained from KOMA Biotech (Seoul, Korea). The generation of anti-PTK7 anti-serum was described previously [57 (link)].

Shin W.S., Park M.K., Kim J.H., Oh S.W., Jang J.Y., Lee H, & Lee S.T. (2022). PTK7, a Catalytically Inactive Receptor Tyrosine Kinase, Increases Oncogenic Phenotypes in Xenograft Tumors of Esophageal Squamous Cell Carcinoma KYSE-30 Cells. International Journal of Molecular Sciences, 23(4), 2391.

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