BD PhosflowTM Alexa Fluor 647 Mouse anti-Pyk2 (pY402)

Description

The specificity of mAb L68-1256.272 was confirmed by western blot analysis using unconjugated antibody on lysates from PBMC that were untreated lane 1 or stimulated by cross-linking of CD3 and CD28 lane 2 . Pyk2 pY402 is identified as a band of 116 kDa, with increased intensity in the stimulated cells. Analysis of Pyk2 pY402 in human peripheral blood lymphocytes. Human peripheral blood mononuclear cells PBMC were either untreated left panel or stimulated by cross-linking of CD3 and CD28 with NA/LE Mouse anti-Human CD3 mAb UCHT1 Cat. No. 555329 and NA/LE Mouse anti-Human CD28 mAb CD28.2 Cat. No. 555725 on ice for 15 minutes, followed by Purified Goat anti-Mouse Ig Cat. No. 553998 on ice for 15 minutes, and then allowed to undergo phosphorylation at 37C for 5 minutes right panel . The cells were fixed BD Cytofix buffer, Cat. No. 554655 for 10 minutes at 37 C, permeabilized BD Phosflow Perm Buffer III, Cat. No. 558050 on ice for 30 minutes, blocked with normal mouse immunoglobulin, and then stained with Alexa Fluor 647 Mouse anti-Pyk2 pY402, Cat. No. 560256 and PE Mouse anti-human CD3 mAb SK7 Cat. No. 347347 . For data analysis, lymphocytes were selected by their scatter profile. The data demonstrates that the upregulated phosphorylation of Pyk2 was restricted to the CD3-positive T lymphocytes under these stimulation conditions. Flow cytometry was performed on a BD FACSCalibur flow cytometry system. The specificity of mAb L68-1256.272 was confirmed by western blot analysis using unconjugated antibody on lysates from PBMC that were untreated lane 1 or stimulated by cross-linking of CD3 and CD28 lane 2 . Pyk2 pY402 is identified as a band of 116 kDa, with increased intensity in the stimulated cells. Show More Analysis of Pyk2 pY402 in human peripheral blood lymphocytes. Human peripheral blood mononuclear cells PBMC were either untreated left panel or stimulated by cross-linking of CD3 and CD28 with NA/LE Mouse anti-Human CD3 mAb UCHT1 Cat. No. 555329 and NA/LE Mouse anti-Human CD28 mAb CD28.2 Cat. No. 555725 on ice for 15 minutes, followed by Purified Goat anti-Mouse Ig Cat. No. 553998 on ice for 15 minutes, and then allowed to undergo phosphorylation at 37C for 5 minutes right panel . The cells were fixed BD Cytofix buffer, Cat. No. 554655 for 10 minutes at 37 C, permeabilized BD Phosflow Perm Buffer III, Cat. No. 558050 on ice for 30 minutes, blocked with normal mouse immunoglobulin, and then stained with Alexa Fluor 647 Mouse anti-Pyk2 pY402, Cat. No. 560256 and PE Mouse anti-human CD3 mAb SK7 Cat. No. 347347 . For data analysis, lymphocytes were selected by their scatter profile. The data demonstrates that the upregulated phosphorylation of Pyk2 was restricted to the CD3-positive T lymphocytes under these stimulation conditions. Flow cytometry was performed on a BD FACSCalibur flow cytometry system. Show More The specificity of mAb L68-1256.272 was confirmed by western blot analysis using unconjugated antibody on lysates from PBMC that were untreated lane 1 or stimulated by cross-linking of CD3 and CD28 lane 2 . Pyk2 pY402 is identified as a band of 116 kDa, with increased intensity in the stimulated cells. Analysis of Pyk2 pY402 in human peripheral blood lymphocytes. Human peripheral blood mononuclear cells PBMC were either untreated left panel or stimulated by cross-linking of CD3 and CD28 with NA/LE Mouse anti-Human CD3 mAb UCHT1 Cat. No. 555329 and NA/LE Mouse anti-Human CD28 mAb CD28.2 Cat. No. 555725 on ice for 15 minutes, followed by Purified Goat anti-Mouse Ig Cat. No. 553998 on ice for 15 minutes, and then allowed to undergo phosphorylation at 37C for 5 minutes right panel . The cells were fixed BD Cytofix buffer, Cat. No. 554655 for 10 minutes at 37 C, permeabilized BD Phosflow Perm Buffer III, Cat. No. 558050 on ice for 30 minutes, blocked with normal mouse immunoglobulin, and then stained with Alexa Fluor 647 Mouse anti-Pyk2 pY402, Cat. No. 560256 and PE Mouse anti-human CD3 mAb SK7 Cat. No. 347347 . For data analysis, lymphocytes were selected by their scatter profile. The data demonstrates that the upregulated phosphorylation of Pyk2 was restricted to the CD3-positive T lymphocytes under these stimulation conditions. Flow cytometry was performed on a BD FACSCalibur flow cytometry system.