GSH/GSSG Ratio Detection Assay Kit (Colorimetric)

Summary & Explanation

GSSG in the samples treated with TCA solution was
reduced with reagents including NaBH4 and NaOH to form
GSH. NaOH was used to increase the pH, which was low
due to the utilization of TCA solution, to enable the reduction
of the GSSG molecules. NaBH4 was used as a reductant.
After the reduction was completed, HCl solution was
added to remove the remnant NaBH4 in order to prevent
extra-reduction of DTNB molecules and re-oxidation of
GSH molecules. GSH levels were measured using the
Ellman method that was modified by Hu [13] using 500 mM
Tris solution (pH: 8.2). In this method, thiol residues of
GSH reduced the DTNB molecules to 2-nitro-5-benzoic
acid which has an absorbance at 412 nm spectrophotometrically.
GSH was measured before and after the reduction
process. After dilution factor multiplication, the GSH
content was subtracted from the total GSH (GSH + GSSG)
content and divided by two equals to the GSSG amount.