Description
Glucose-6-Phosphate Assay Kit (BA0111) (BA0111)
The Glucose-6-Phosphate Assay Kit (SKU: BA0111) provides a simple, automation-ready colorimetric method for measuring glucose-6-phosphate (G6P). Most glucose entering cells is phosphorylated to G6P, which lies at the start of glycolysis and the pentose phosphate pathway and can also be stored as glycogen. In the assay, G6P is oxidised by glucose-6-phosphate dehydrogenase and the NADPH formed is coupled to the WST-8 formazan chromogen. The intensity of the product colour, measured at 460 nm, is proportional to the G6P concentration in the sample. The homogeneous mix-incubate-measure format is readily automated for high-throughput screening.
| Product Name: | Glucose-6-Phosphate Assay Kit (BA0111) |
| SKU: | BA0111 |
| Detection Method: | Colorimetric (460 nm) |
| Detection Range: | 10 - 1000 uM G6P |
| Sample Type: | Plasma, serum, tissue and culture media |
| Species Reactivity: | All |
| Assay Time: | 20 minutes |
| Kit Size: | 100 Assays |
| Equipment Required: | Microplate reader |
| Storage: | -20C |
| Shelf Life: | 6 months after receipt |
| Shipping: | Gel Pack |
A colorimetric assay for glucose-6-phosphate in biological samples using a WST-8 coupled detection system.
- Fast and sensitive with a linear detection range of 10 - 1000 uM G6P
- Convenient, high-throughput homogeneous mix-incubate-measure format
- Readily automated for high-throughput screening
- G6P determination in biological samples such as plasma, serum, tissue and culture media
Note: The below protocol is a sample protocol. Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.
| Step | Procedure |
| 1 | Sample preparation: homogenise tissue or ~2 x 10^6 cells in 100 uL PBS and centrifuge at 14,000 rpm for 5 min; use clear supernatant. Serum and plasma can be measured directly but may need a sample blank if they absorb at 460 nm. |
| 2 | Standards: dilute the 100 mM Standard to 1000 uM Premix by mixing 5 uL with 495 uL distilled water, then dilute per the table. Transfer 20 uL of each standard into a 96-well plate. |
| 3 | Samples: add 20 uL of each sample to separate wells. For samples with background absorbance at 460 nm or high NADH/NADPH, add a second aliquot as a sample blank. |
| 4 | Prepare Working Reagent per reaction from 75 uL Assay Buffer, 8 uL NADP/WST8, 1 uL Enzyme A and 1 uL Enzyme B (omit Enzyme A for a blank working reagent). Add 80 uL to each standard and sample well, mix and incubate protected from light for 20 min at room temperature. |
| 5 | Read OD460nm. |
Subtract the blank value from the standard values and plot delta-OD against standard concentration. [G6P] = (ODSAMPLE - ODBLANK) / Slope x n (uM), where ODBLANK is standard 4 or the sample blank and n is the dilution factor. If the calculated concentration exceeds 1000 uM, dilute in distilled water and repeat. 100 uM G6P equals 34 mg/L, 0.0034% or 34 ppm.
| Component | Quantity | Storage |
| Assay Buffer | 10 mL | -20C |
| NADP/WST8 | 1 mL | -20C |
| Standard (100 mM G6P) | 100 uL | -20C |
| Enzyme A | 120 uL | -20C |
| Enzyme B | 120 uL | -20C |
