Method for detecting activity of spermidine/spermine N1-acetyltransferase
A technology of acetyltransferase and activity, applied in the field of detection of spermidine/spermine N1-acetyltransferase activity, which can solve problems such as inappropriate, rapid heart rate and slow action
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Embodiment 1
[0363] The liver cells of 20 rats (Wistar rats, female, 200-300g) were used, and the cells were obtained by Celsis in vitro technology and stored in liquid nitrogen. The day before use, the frozen hepatocytes were thawed in a 37°C water bath, suspended with the culture medium, centrifuged at 100×g for 10 minutes, and resuspended with Krebs-Hensleite buffer (KHB). Use 24-well plates for in vitro culture at 37°C with 5% CO 2 , 95% air, each hole is filled with about 2.5×10 5 Live liver cell stock solution 0.49ml, incubated for 10min. Then pre-incubation for 10min. In the liver cell solution, add L-dopa at 200 μM, inhibitor quercetin at 480 μM, add L-dopa at 200 μM, inhibitor methotrexate at 50 μM, and add L-dopa at 200 μM, inhibitor acetamine Phenol 2144μM, each substrate, inhibitor mixed solution 5μl to the culture plate, continue to incubate for 0, 30 and 60min. At the end of the incubation, 0.5 ml of ice methanol was added to each well to stop the reaction. Each group wa...
Embodiment 2
[0367] The liver cells of 20 rats (Wistar rats, female, 200-300 g) were used, and the cells were obtained by Celsis in vitro technology and stored in liquid nitrogen. The day before use, the frozen hepatocytes were thawed in a 37°C water bath, suspended with the culture medium, centrifuged at 100×g for 10 minutes, and resuspended with Krebs-Hensleite buffer (KHB). Use 24-well plates for in vitro culture at 37°C with 5% CO 2 , 95% air, each hole is filled with about 2.5×10 5 Live liver cell stock solution 0.49ml, incubated for 10min. Then pre-incubation for 10min. In the liver cell solution, add L-dopa at 20 μM, inhibitor quercetin at 480 μM, add L-dopa at 200 μM, inhibitor methotrexate at 50 μM, and add L-dopa at 200 μM, inhibitor acetamine Phenol 2144μM, each substrate, inhibitor mixed solution 5μl to the culture plate, continue to incubate for 0, 30 and 60min. At the end of the incubation, 0.5 ml of ice methanol was added to each well to stop the reaction. Each group wa...
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