Hello you all,
we want to perform an Annexin V and PI staining on our B cell culture, does anybody know if using the commercial buffer (biolegend in our case) is absolutely necessary? Or can we substitue it by standard in house buffers (supplemented with Ca2+, i guess)?
Thanks for your help, Caro
hi
Typically, the combinations of buffering kits are Patented for each company. also Annexin V and PI in your kit is ready to use .
1- PI dye Usually provided in DDW at 50mg/ml.
2- The Annexin V Binding Buffer is a 10X concentrate composed of a 0.2 µm sterile filtered 0.1M Hepes (pH 7.4), 1.4M NaCl, and 25 mM CaCl2 solution. Prior to staining cells, an appropriate quantity of a 1X working solution should be made by diluting the 10X concentrate 1:10 with distilled water.
good luck
The Annexin V Binding Buffer is a 10X concentrate composed of a 0.2 µm sterile filtered 0.1M Hepes (pH 7.4), 1.4M NaCl, and 25 mM CaCl2 solution.
In the presence of calcium, Annexin V binds to phosphatidylserine allowing apoptotic cells to be easily identified by flow cytometry or fluorescence microscopy. Therefore, without CaCl2 binding affinity will reduce.
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