Hello.
I am new to the GSH assay and I am learning about it online. I performed it this week for the first time.
My protocol said to add DTNB and GR to my GSH standards + samples - allow 30 sec the conversion of GSSG to GSH - and THEN - add NADPH and measure absorbance.
Is not NADPH essential for the conversion of GSSG to GSH? Why adding it later?
Also. Can I add just DTNB and measure the absorbance (initial GSH) and then add GR+NADPH and measure absorbance again to measure the total GSH (Initial GSH + reduced GSSG)?
So I can calculate also GSSG (GSHt - GSHi) with this assay (without using vinylipyridine) ?