Joan Years ago I looked at a potential flow method for glutathione peroxidase. This is based on the rate of loss of GSH in cells when glutathione reductase is inhibited. We used the nitrosourea BCNU (a chemotherapy drug) to inhibit glutathione reductase, based on earlier work done in Jim Watson's lab (I believe part of Caroline Dive's PhD thesis). We measured GSH using monobromobimane. We found that glutathione levels decreased with time at 37°, presumably under the action of glutathione peroxidase. Therefore a plot of cellular GSH vs. time following BCNU treatment would probably give a rough approximation of glutathione peroxidase activity. However, this area of cellular biochemistry is very tricky and you very definitely would want an external biochemical standard of enzyme activity in order to convince yourself that the technique was valid. If you try this approach, I would be very interested to hear how you get on. I never really finished the project because at that time we didn't have the flow GSH methodology working well enough. David Hedley Ontario Cancer Institute Toronto. > -----Original Message----- > From: joan Kalnitsky [SMTP:jkalnits@vt.edu] > Sent: Wednesday, September 26, 2001 9:17 AM > To: Cytometry Mailing List > Subject: glutathione peroxidase > > > > I have a client who has been measuring glutathione peroxidase in > white > blood cells using a non flow method. He would very much like to be able > to > look at the difference in response of the lymphocyte vs phagocytic cell > populations and has inquired about the possibility of using flow , so we > could simultaneously look at both individual populations. > I do not have a protocol for measuring (GSHPX) glutathione > peroxidase. I > did check the archives and did not find anything of great help. Can > anyone > suggest a method/protocol that would help me help him achieve his goal. > Thank you in advance, > Joan K. > > > Flow Cytometry Lab Supervisor > VMRCVM > (540) 231-4115 > FAX 540-231-7367 > jkalnits@vt.edu > > "It is better to serve than to receive." > B. Borg
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