Rabbit anti lamin b1

Normal and DOX-induced senescent 293 ​T cells were fixed with 4% paraformaldehyde (PFA) for 15 ​min, permeabilized with 0.2% Triton X-100 for 3 ​min and was then blocked with 10% (v/v) BSA (bovine serum albumin) in PBS for 2 ​h at room temperature or at 4 ​°C overnight. Subsequently, the cells were washed with PBS three times and were then incubated with primary antibodies rabbit anti-Forkhead box O4 protein (FOXO4) (1:150, Proteintech, U.S.A), rabbit anti-p16^ink4a (1:150, Abclonal, China), or rabbit anti-lamin B1 (1:150, Abclonal, China) for 2 ​h. After the incubation, the cells were washed three times with PBS and incubated with goat anti-rabbit IgG H&L Alexa Fluor® 488 (1:200, Abcam, UK) for 2 ​h. Then, after three washes with PBS, the cells were stained with DAPI for 5 ​min, followed by another three washes with PBS. Finally, the cells were mounted and imaged on an inverted fluorescence microscope (Zeiss, Germany). The fluorescence intensity was obtained from 3 randomly selected regions of the cell culture to determine the expression levels of FOXO4, p16^ink4a, and lamin B1 in each group. Three independent batches of cells were analyzed to determine the significance of difference between different groups.

Mice were anesthetized and perfused with PBS. The kidneys were collected, fixed with 4% PFA, and cut to obtain 30 ​μm sections using a vibratome (Invitrogen, U.S.A). After three rinses in PBS (5 ​min each), the sections were blocked in PBST (PBS ​+ ​0.3% Triton X-100) with 3% BSA for 2 ​h, and were then incubated in primary antibodies rabbit anti-lamin B1 (Abclonal, China), mouse anti-p53 (Abcam, UK) or rabbit anti-p21 (ABclonal, China) with a dilution of 1:1000 ​at 4 ​°C for 24 ​h, followed by three rinses in PBS. Then, the sections were incubated with goat anti-rabbit IgG H&L Alexa Fluor® 488 (1:300, Abcam, UK) or goat anti-mouse IgG H&L Alexa Fluor® 488 secondary antibodies (1:300, Abcam, UK) for 2 ​h at room temperature. After the incubation, the sections were washed with PBS for 5 ​min, and the nuclei were counterstained with DAPI for 5 ​min. Finally, the sections were washed in PBST and mounted in mounting medium. Fluorescence images were acquired with a 20 ​× ​objective on a confocal microscope.
Flies were anesthetized with CO₂, dissected in PBS, and fixed with 4% PFA at room temperature. Brain sections were prepared and incubated with rabbit anti-caspase-3 primary antibody (1:500, Beyotime, China) and then with goat anti-rabbit IgG H&L Alexa Fluor® 488 (1:200, Abcam, UK), followed by DAPI staining and fluorescence imaging.