rabbit anti β galactosidase  (Valiant Co Ltd)

Journal: Scientific Reports

Article Title: Numb mitigates intestinal epithelial cell senescence induced by radiation through a PLK1-dependent pathway

doi: 10.1038/s41598-026-44793-x

Figure Lengend Snippet: Inhibition of Numb expression enhanced radiation-induced senescence in intestinal epithelial cells. (A-C) Immunohistochemical staining of intestinal sections and western blotting using intestinal mucosal extracts to assess active caspase-3, γH2AX, and p21 expression on days 3 and 7 after 13 Gy WAI. Representative images are displayed. Scale bar = 100 μm. Quantitative analysis of the western blotting results are presented in the form of statistical bars on the right panel of the corresponding immunohistochemistry images. n = 3 mice in each group. (D) Frozen sections of the intestine were stained for senescence-associated β-galactosidase (SA-β-gal) activity in intestinal epithelial cells on days 3 and 7 following 13 Gy WAI. SA- β-galactosidase levels were quantified via western blotting using intestinal mucosal extracts and are presented on the right panel. n = 3 mice in each group. The values are presented as the means ± SEMs. ** P < 0.01, * P < 0.05.

Article Snippet: After blocking with TBST containing 5% nonfat dry milk, specific primary antibodies against Numb (Abcam, Ab220362 ), p53 (Abcam, Ab131442 ), p21 (Santa Cruz, sc-377515), p16 (Abcam, ab211542), γH2AX (Santa Cruz, sc-517348), Cyclin B (Santa Cruz, sc-166210), Plk1 (Abcam, ab12157) and β-galactosidase (CST, #27198) were incubated with the membranes at 4 °C overnight, followed by incubation with the respective secondary antibodies.

Techniques: Inhibition, Expressing, Immunohistochemical staining, Staining, Western Blot, Immunohistochemistry, Activity Assay

Journal: Scientific Reports

Article Title: Numb mitigates intestinal epithelial cell senescence induced by radiation through a PLK1-dependent pathway

doi: 10.1038/s41598-026-44793-x

Figure Lengend Snippet: Inhibition of Numb promoted mitotic arrest and cellular senescence after radiation in colonic cells. (A) Flow cytometry was utilized to assess the apoptosis of HCT116 cells following 24 h and 72 h of radiation. n = 3 repeats in each group. (B) Flow cytometry was utilized to analyze the cell cycle distribution of colonic HCT116 cells at 24 h and 72 h postradiation. ** P < 0.01 IR versus IR+shNumb group at 24 h, * P < 0.05 IR versus IR+shNumb group at 72 h. n = 3 repeats in each group. (C) Immunofluorescence staining was conducted to evaluate the expression of the DNA damage marker γH2AX after 72 h of radiation. Scale bar = 50 μm. (D) Cell senescence-associated β-gal staining was performed to analyze cell senescence after 72 h of radiation. Scale bar = 50 μm.

Article Snippet: After blocking with TBST containing 5% nonfat dry milk, specific primary antibodies against Numb (Abcam, Ab220362 ), p53 (Abcam, Ab131442 ), p21 (Santa Cruz, sc-377515), p16 (Abcam, ab211542), γH2AX (Santa Cruz, sc-517348), Cyclin B (Santa Cruz, sc-166210), Plk1 (Abcam, ab12157) and β-galactosidase (CST, #27198) were incubated with the membranes at 4 °C overnight, followed by incubation with the respective secondary antibodies.

Techniques: Inhibition, Flow Cytometry, Immunofluorescence, Staining, Expressing, Marker

Journal: Scientific Reports

Article Title: Numb mitigates intestinal epithelial cell senescence induced by radiation through a PLK1-dependent pathway

doi: 10.1038/s41598-026-44793-x

Figure Lengend Snippet: PLK1 overexpression compensated for the Numb knockdown-induced cellular senescence phenotype in colonic cells after radiation. (A) Senescence-associated β-gal staining was performed on different groups of HCT116 cells. The results of the quantitative analysis of the percentage of cells with positive β-gal staining are shown in the right panel. Scale bar = 50 μm. (B) Colony formation assay illustrating colonies in different groups of HCT116 cells after radiation. Quantitative analysis of the total optical density of the colony formation assay is shown in the right panel. (C-E) RT‒PCR was used to detect the expression of the indicated cytokines (IL-6, IL-8, and IL-1β) in different groups of cells. The expression levels were normalized to those of β-actin. The values are presented as the means ± SEMs. ** P < 0.01, * P < 0.05. n = 3 repeats in each group.

Article Snippet: After blocking with TBST containing 5% nonfat dry milk, specific primary antibodies against Numb (Abcam, Ab220362 ), p53 (Abcam, Ab131442 ), p21 (Santa Cruz, sc-377515), p16 (Abcam, ab211542), γH2AX (Santa Cruz, sc-517348), Cyclin B (Santa Cruz, sc-166210), Plk1 (Abcam, ab12157) and β-galactosidase (CST, #27198) were incubated with the membranes at 4 °C overnight, followed by incubation with the respective secondary antibodies.

Techniques: Over Expression, Knockdown, Staining, Colony Assay, Expressing