I added fructose 1,6 phosphate into cell culture medium and did the cck8 assay. The group with fructose 1,6 phosphate have significant higher absorption at 450nm, while under microscope I did not find significant change of cell number? Does fructose 1,6 phosphate interfere with cck8 assay? What is the mechanism?
This kit reaction is based in NADH and NADPH reactivity (reducing power is a marker of viability). You are boosting the central metabolism with an energy rich molecule fructose 1,6 phosphate (http://www.ncbi.nlm.nih.gov/books/NBK22423/). This increases the amount of NADH and NADPH and in turn enhance the chromogenic reaction with your kit (http://www.dojindo.com/store/p/456-Cell-Counting-Kit-8.html). To compare the viability of two cultures, the metabolic state of NADH and NADPH should be similar (a limitation of the kit that the manufacturer does not tell you). I would change to FACS or classic trypam blue of possible. Hope this help.
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