Glucose Uptake-Glo™ Assay Technical Manual
Instructions for Use of Product(s)
J1341, J1342, J1343
Literature # TM467
The Glucose Uptake-Glo™ Assay is a nonradioactive, plate-based, homogeneous bioluminescent method for measuring glucose uptake in mammalian cells based on the detection of 2-deoxyglucose-6-phosphate (2DG6P). When 2-deoxyglucose (2DG) is added to cells, it is transported across the membrane and rapidly phosphorylated in the same manner as glucose. However, enzymes that further modify glucose-6-phosphate (G6P) cannot modify 2DG6P, and thus this membrane-impermeable analyte accumulates in the cell. After a brief period of incubation, an acid detergent solution (Stop Buffer) is added to lyse cells, terminate uptake and destroy any NADPH within the cells. A high-pH buffer solution (Neutralization Buffer) is then added to neutralize the acid. A Detection Reagent containing glucose-6-phosphate dehydrogenase (G6PDH), NADP+, Reductase, Ultra-Glo™ Recombinant Luciferase and proluciferin substrate is added to the sample wells. G6PDH oxidizes 2DG6P to 6-phosphodeoxygluconate and simultaneously reduces NADP+ to NADPH. The Reductase uses NADPH to convert the proluciferin to luciferin, which is then used by Ultra-Glo™ Recombinant Luciferase to produce a luminescent signal that is proportional to the concentration of 2DG6P.
Summary of Changes
The following changes were made to the 4/21 revision of this document:
1. The Table of Contents was updated.
2. Section 3.E was added.
3. The numbering of steps in Section 5.D was corrected.
4. The cover was updated.
Revised 4/21.