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HBSS replacement for washing steps in antioxidant assay using flow cytometry


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#1 yean_ny_nie

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Posted 19 August 2010 - 01:18 AM

Hi all,

I'm planning to do a FACS analysis on antioxidant properties in certain protein (which is already transfected inside cancer cells) using flow cytometry. From the protocol that I can get, it requires me to wash and resuspended the cells with HBSS (Hank's balanced salt solution). I'm wondering if I could replace the HBSS with PBS. However, I'm thinking if it will affect the reading upon replacement with PBS. Anyone has encounter this before? Your comments and suggestions is very much welcome.

Thanks a lot for your feedback and best of regards.

#2 Piersgb

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Posted 19 August 2010 - 01:30 AM

The change in buffer may have some effect on your flow results, the best way to find out is to compare the results of the same stainings with different buffers yourself with your samples. The difference between PBS and HBSS is shown below:


Google: "HBSS PBS recipe" :

HBSS
400 ml H2O

4.0 g NaCl (=13.7 ml 5 M NaCl)
0.20 g KCl
0.50 g Glucose
0.021 g Na2HPO4
0.027 g KH2PO4
0.096 g CaCl2
0.111 g MgCl2
0.176 g NaHCO3

qc 500 ml
pH 7.1
Filter sterilize

PBS
900 ml H2O
8 g NaCl or 27.4 ml 5M
0.2 g KCl
0.19 g KH2PO4 or 1.4 ml 1M
0.61 g Na2HPO4 or 4.3 ml 1M

qc 1 L
pH 7.4




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