P21 rabbit polyclonal antibody

Three samples at PN5 were included, and at least three slices with good morphology from each sample were selected for immunofluorescence analysis. The procedures before primary antibody incubation were the same as the methods of immunohistochemistry mentioned above. The slices were incubated overnight (15 h) with diluted KLF6 mouse monoclonal antibodies (1:50, Santa Cruz, # sc-7158) and then treated with FITC conjugated goat anti-mouse IgG (Boster, #BA1101). Subsequently, the slices were treated with goat serum, following which the slices were incubated overnight (15 h) with diluted P21 rabbit polyclonal antibody (1:100, Proteintech, # 28248-1-AP). Wash four times with PBS. The slices were treated with Cy3 conjugated goat anti-rabbit IgG (Boster, #BA1032) in an immunohistochemical wet box. All the slices were counterstained with DAPI dye (Beyotime, #C1006)) for 5 min away from light. A Leica SP8 confocal laser scanning microscope was used to observe the fluorescence intensity. All slices were prepared under the same conditions and time to ensure homogeneity between samples. Images were taken under identical gain and offset values, with the same intensity of excitation wavelength and exposure time.

The culture medium was removed, and after washing the cells twice with PBS, they were harvested for further analysis. The preparation of cell lysates and Western blotting were performed by the method of Sun et al. [42 (link)]. Antibodies used were p16 INK4A (F-12) (Santa Cruz Biotechnology, sc-1661), P53 mouse monoclonal antibody (Proteintech, 60283-2-Ig), P21 rabbit polyclonal antibody (Proteintech, 27296-1-AP), LC3B antibody (Cell Signaling, 2575S), LAMP2 (Abcam, ab13524), AMPK (Cell Signaling, D63G4), P-AMPK (Cell Signaling, 2535S), p70 S6 Kinase Antibody (Cell Signaling, 9202S), p-p70 S6 Kinase Antibody (Cell Signaling, 9234S), and GAPDH polyclonal antibody (Proteintech, 10494-1-AP).