Image analysis equipment

Full‐length ApoE, ApoE fragments, and Tau5, AT8, and PHF1 expressions in the hippocampus was detected using western blotting. Using a 4–20% double TIS polyacrylamide gel (Cat#M00655, Gen Script Biotech Corp., Nanjing, China) or a standard XT 4–12% double TIS gel (Bio‐Rad, USA), the samples were separated from SDS‐PAGE and transferred to nitrocellulose membranes (Bio‐Rad, USA). The membranes were then blocked with 5% skimmed milk powder and identified using the following primary antibodies: Tau5 (1:1000, Cat#ABN454, Millipore, USA), full‐length ApoE and ApoE fragments (1:4000, Cat#178479, Calbiochem, Germany), PHF1 (1:1000, Cat#3Ab184951, Abcam, UK), and AT8 (1:2000, Cat#MN1020, Thermo Fisher Scientific, USA). After washing at 4°C overnight, the membranes were incubated at 37°C for 1 h with goat anti‐mouse antibody (1:5000, Cat#31430, Invitrogen, USA). The nitrocellulose membranes were then scanned and photographed using Bio‐Rad image analysis equipment after being washed in TBST and dripped with ECL chemiluminescent liquid (Cat#34577, Invitrogen, USA). The ratio of the integral optical density of the target band to that of the β‐actin band represents the level of expression of the target protein. A 100% change refers to control or sevoflurane levels for comparison between the experimental settings.

50 µg of protein was then separated on 10% sodium dodecyl sulfate-polyacrylamide gel and transferred to polyvinylidene fluoride membranes (Millipore). After 2 h of blovarian carcinomaking in nonfat milk, membranes were incubated overnight at 4°C with primary antibodies against TGF1B (1 : 1,000, ab96099, Abcam), Smad2/3 (1 : 5,000, ab179461, Abcam), p-Smad2/3 (1 : 5,000, ab124956, Abcam), and β-actin (1 : 5,000, ab8227, Abcam) (1 : 20,000, ab6721, Abcam). Utilizing a Bio-Rad Image analysis equipment, enhanced chemiluminescence reagent was employed to evaluate protein blots (Bio-Rad). The relative protein level (relative to -actin) was assessed using ImageJ.