Total proteins were extracted using RIPA lysis buffer. Western blot was used to detect the expression of POP, MMP2, β-actin, and TGF-β1. The total protein was separated on 12% sodium dodecyl sulfate–polyacrylamide gel electrophoresis gels and transferred to the PVDF membrane. The blot was blocked with 5% skim milk solution for 2 h at room temperature and was incubated with primary POP antibody (dilution 1: 1000, BS60084, BIOWORLD), TGF-β1 (dilution 1:500, BA2120, BOSTER, Wuhan, China), MMP2 (dilution 1:1000, BA2120, BOSTER), and β-actin (dilution 1:5000, 66009-1-Ig, Proteintech, Rosemont, IL, USA) overnight at 4 °C. After washing three times with TBST, the blot was incubated with anti-mouse IgG (dilution 1:20,000, BS12478, BIOWORLD) or anti-rabbit IgG (dilution 1:20,000, BS13278, BIOWORLD) for 1 h at room temperature. An Enhanced Easy See Western Blot Kit was employed in order to visualize the target bands, and the intensity of bands was quantified by Bio-Rad Image Laboratory software (Version 6.0). β-actin was used as an internal reference for detecting relative expression levels.
Han S., Wang S., Fan X., Chen M., Wang X., Huang Y., Zhang H., Ma Y., Wang J, & Zhang C. (2023). Abnormal Expression of Prolyl Oligopeptidase (POP) and Its Catalytic Products Ac-SDKP Contributes to the Ovarian Fibrosis Change in Polycystic Ovary Syndrome (PCOS) Mice. Biomedicines, 11(7), 1927.
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