Regulating the level of septin2 sulfhydryl nitrosylation modification for pharmaceutical use
A technology of lv-septin2c111a and a drug is applied in the field of pharmaceutical use for regulating the level of sulfhydryl nitrosylation of Septin2, so as to achieve the effect of inhibiting the occurrence of aortic aneurysm and aortic dissection
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[0018] 1. Biotin-Switch method to detect sulfhydryl nitrosylation modified proteins (S-nitrosylation Protein Detection Kit from Cayman Chemical Company, Cat. No. 10006518)
[0019] (1) After the cells are treated, remove the culture medium, wash with pre-cooled wash buffer 3 times, add 500 μL wash buffer to scrape off, and transfer to a 1.5 mL centrifuge tube.
[0020] (2) Centrifuge at 500Xg at 4°C for 5 minutes, discard the supernatant.
[0021] (3) Place the cells on ice and configure Blocking buffer. Specific preparation method: Add new blocking reagent to 100 μL DMF and 900 μL buffer A to make the stock solution of blocking buffer. After mixing, add 9 mL of buffer A to obtain the blocking buffer.
[0022] (4) Add 500 μL blocking buffer to each tube to resuspend the cell pellet obtained in (2), and rotate at 4°C for 30 minutes.
[0023] (5) Centrifuge at 12,000 rpm at 4°C for 10 min, and transfer the supernatant to a 15 mL centrifuge tube.
[0024] (6) Add 4 times the vo...
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