Antibodies for flow cytometry were purchased from Cell Signaling Technologies (pY397FAK #3283, pPaxilin #2541), abcam (TET3 #ab135033, 5hmU #ab19735), Novus Biologicals (laminin #NB300–144), Active Motif (5mC #39649) or BD Biosciences (pSTAT3 #557815). Secondary antibodies used were coupled to either Alexa Fluor 647 or Alexa Fluor 488 (Invitrogen) diluted up to 1:4,000 in PBS/1% BSA staining buffer. For blocking FAK activity, GSC cell lines were treated with 10 μM FAK inhibitor (Santa Cruz #sc-203950). Intracellular staining was performed after fixation of single cell suspensions with 2% paraformaldehyde and permeabilization with ice-cold 100% methanol, blocked with PBS/1% BSA for 1h at 4°C, and stained with an antibody diluted at 1:50 in PBS/1% BSA for 30 – 45 min at room temperature. Detection of DNA modification was performed upon RNA digestion using RNase A at 40 μg/ml for 10 min at 37°C prior to blocking procedure. Stained single cell suspensions were analyzed using an AccuriC6 cytometer (BD Biosciences), followed by FlowJo 7.6.1 analysis.
Pstat3 557815
Manufactured by BD
The PSTAT3 #557815 is a laboratory equipment product manufactured by BD. It is designed to perform specific functions within a research or clinical laboratory setting. The core function of this product is to provide essential capabilities for laboratory tasks, but a detailed description cannot be provided while maintaining an unbiased and factual approach.
Automatically generated - may contain errors
2 protocols using pstat3 557815
1
Antibody-based Flow Cytometry Analysis
2
Antibody-based Flow Cytometry Analysis
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Antibodies for flow cytometry were purchased from Cell Signaling Technologies (pY397FAK #3283, pPaxilin #2541), abcam (TET3 #ab135033, 5hmU #ab19735), Novus Biologicals (laminin #NB300–144), Active Motif (5mC #39649) or BD Biosciences (pSTAT3 #557815). Secondary antibodies used were coupled to either Alexa Fluor 647 or Alexa Fluor 488 (Invitrogen) diluted up to 1:4,000 in PBS/1% BSA staining buffer. For blocking FAK activity, GSC cell lines were treated with 10 μM FAK inhibitor (Santa Cruz #sc-203950). Intracellular staining was performed after fixation of single cell suspensions with 2% paraformaldehyde and permeabilization with ice-cold 100% methanol, blocked with PBS/1% BSA for 1h at 4°C, and stained with an antibody diluted at 1:50 in PBS/1% BSA for 30 – 45 min at room temperature. Detection of DNA modification was performed upon RNA digestion using RNase A at 40 μg/ml for 10 min at 37°C prior to blocking procedure. Stained single cell suspensions were analyzed using an AccuriC6 cytometer (BD Biosciences), followed by FlowJo 7.6.1 analysis.
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