Enhanced chemiluminescence western blot detection kits

Western blot analysis was conducted as previously described (Meng et al., 2014 (link)). Right cortex tissues were collected from each rat (n = 3), and the total protein was extracted using a protein extraction reagent supplemented with protease and phosphatase inhibitor cocktails (ComWin Biotech, China). The total protein concentration of each sample was determined by a BCA kit (ComWin Biotech, China). Equal amounts of protein were separated using SDS-polyacrylamide gel electrophoresis and were transferred to nitrocellulose membranes. These membranes were blocked before being incubated overnight at 4°C with the appropriate primary antibodies: cnpase (ab183500, 1:1000), MBP (ab209328, 1:1000), Vimentin (ab92547, 1:10,000), SYP (ab32127, 1:1000), PSD95 (ab76115, 1:1000), MAP-2 (ab32454, 1:2000), Tau-1 (ab75714, 1:1000), BDNF (ab108319, 1:1000), p-TrkB (Tyr705) (ab229908, 1:1000), TrkB (CST4603, 1:1000), p-CREB (Ser133) (ab32096, 1:1000), CREB (ab32515, 1:1000), p-Akt (Ser473) (CST4060, 1:1000), Akt (CST4685, 1:1000), and β-actin (EXP0036 F, 1:2000). Then, the membranes were washed three times and incubated with the appropriate secondary antibodies. The blots were visualized using enhanced chemiluminescence western blot detection kits (ComWin Biotech, China). The blot densities were calculated by ImageJ software.

Western blot analysis was conducted as previously described (Xie et al. 2019 ; Zhu et al. 2021d (link)). Right cortex tissues were collected from each rat (n = 3), and the total protein was extracted using a protein extraction reagent supplemented with protease and phosphatase inhibitor cocktails (ComWin Biotech, Beijing, China). The total protein concentration of each sample was determined by a BCA kit (ComWin Biotech, Beijing, China). Equal amounts of protein were separated using SDS-polyacrylamide gel electrophoresis and transferred to nitrocellulose membranes. The membranes were blocked before being incubated overnight at 4 °C with the appropriate primary antibodies against the following proteins: NLRP3 (A5652, 1:1000), IL-1β (Ab16288, 1:1000), Nrf2 (A0674, 1:1000), HO-1 (A1346, 1:1000), Akt (CST4685, 1:1000), p-Akt (CST4060T, 1:1000) and β-actin (EXP0036 F, 1:2000). Then, the membranes were washed three times and incubated with the appropriate secondary antibodies. The blots were visualized using enhanced chemiluminescence western blot detection kits (ComWin Biotech, Beijing, China). The blot densities were measured by Image J software (Bethesda, MD).