Description
GST Activity Assay Kit (Colorimetric) (BA0028) (BA0028)
The GST Activity Assay Kit (Colorimetric) (SKU: BA0028) is a convenient assay for the quantitative determination of glutathione S-transferase (GST) activity in biological samples. GST (EC 2.5.1.18) is a multifunctional enzyme central to cellular detoxification, protecting cells against carcinogens and drugs by catalysing the conjugation of glutathione to the electrophilic or hydrophobic sites of foreign compounds. The assay is based on the GST-catalysed reaction between glutathione (GSH) and the substrate CDNB (1-chloro-2,4-dinitrobenzene), which forms a dinitrophenyl thioether that can be detected spectrophotometrically at 340 nm. The rate of increase in absorbance at 340 nm is directly proportional to GST activity, giving a linear range of 2 to 80 U/L over a ten-minute reaction at 25°C. The homogeneous format is convenient and readily automated for high-throughput screening of cell lysates and tissues.
| Product Name: | GST Activity Assay Kit (Colorimetric) (BA0028) |
| SKU: | BA0028 |
| Detection Method: | Colorimetric kinetic; conjugation of GSH to CDNB measured at 340 nm |
| Detection Range: | 2 to 80 U/L for a 10-minute reaction at 25°C (20 µL sample) |
| Sample Type: | Cell lysates, tissues |
| Species Reactivity: | All |
| Assay Time: | 10 minutes |
| Kit Size: | 100 Assays |
| Equipment Required: | Microplate reader |
| Storage: | Store all components at 4°C upon receipt. |
| Shelf Life: | 13 months after receipt. |
| Shipping: | Room Temperature |
A colorimetric kinetic assay for the quantitative determination of glutathione S-transferase (GST) activity. The assay measures the GST-catalysed conjugation of glutathione (GSH) to CDNB, which forms a product detected at 340 nm; the rate of absorbance increase is directly proportional to GST activity. Linear detection range is 2 to 80 U/L for a 10-minute reaction at 25°C.
- Fast and sensitive: linear detection range 2 to 80 U/L for a 10-minute reaction at 25°C (20 µL sample).
- Convenient and high-throughput: homogeneous mix-incubate-measure format, readily automated on HTS liquid-handling systems.
- Glutathione S-transferase activity determination in biological samples such as cell lysates and tissues.
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 (50 mg) in ~250 µL cold 100 mM potassium phosphate (pH 7.0) with 2 mM EDTA, freeze at -80°C to lyse, thaw and centrifuge at 10,000 g for 15 min at 4°C. Prepare cell lysates (~4 million cells) similarly. Samples may be stored at -20 to -80°C for at least one month. |
| 2 | Reagent preparation: bring all reagents to the reaction temperature. Reconstitute the Glutathione tube with 120 µL dH2O and vortex; unused reconstituted glutathione is stable for three weeks at -20°C. |
| 3 | Transfer 20 µL of each sample into separate wells. |
| 4 | Prepare Working Reagent per 96-well assay by mixing 184 µL Assay Buffer, 1 µL reconstituted Glutathione and 1 µL CDNB. Add 180 µL to all samples and tap briefly to mix. |
| 5 | Read OD340nm at time 0 and at least four further time points between 0 and 10 min; kinetic reading every minute for 10 min is recommended. |
Plot OD340nm versus time and use values in the linear range. GST Activity = (ODt2 - ODt1) / t x (10 / 0.00503 µM-1) x n (U/L), where ODt2 and ODt1 are OD values at two time points, t is the time difference, and n is the sample dilution factor. One unit conjugates 1 µmole of CDNB per minute under the assay conditions.
| Component | Quantity | Storage |
| Assay Buffer | 25 mL | 4°C |
| Glutathione | Powder | 4°C |
| CDNB | 120 µL | 4°C |
