wst 8 assay solution  (Dojindo Labs)

Journal: Cell Death & Disease

Article Title: O -GlcNAcylation of XRCC4 controls its stability and confers resistance to DNA double-strand break damage in cancer cells

doi: 10.1038/s41419-025-08209-4

Figure Lengend Snippet: A , B HCT116 cells were treated with 10 μg/mL bleomycin. Cells were collected with the indicated time. A The cell lysates were immunoblotted with the indicated antibodies. XRCC4 and O -GlcNAcylation levels were normalized to GAPDH levels ( n = 4). B Endogenous O -GlcNAcylation of XRCC4 at the indicated time was detected by immunoblotting with RL2 antibody. O -GlcNAcylated XRCC4 was normalized to immunoprecipitated XRCC4 ( n = 3). C Total RNA was extracted from HCT116 cells treated with or without 10 μg/mL bleomycin for 24 h. XRCC4 mRNA levels were quantified by RT-qPCR and normalized to β-actin levels ( n = 5). D HCT116 XRCC4 KO cells stably expressing WT or T308A XRCC4 were treated with or without 10 μg/mL bleomycin for 24 h. The cell lysates were immunoblotted with the indicated antibodies. XRCC4 expression was normalized to GAPDH levels ( n = 5). E HCT116 XRCC4 knockout (KO) cells stably expressing either wild-type (WT) or T308A mutant XRCC4 were treated with 10 μg/mL bleomycin. Cells were harvested at the indicated time points, and whole-cell lysates were subjected to immunoblotting with the specified antibodies. XRCC4 and γH2AX expression levels were normalized to GAPDH. The bar graph depicts relative γH2AX levels at 24 and 48 h in HCT116 XRCC4 KO cells stably expressing WT or T308A XRCC4 ( n = 3). F XRCC4 was knocked down using siRNA and treated with or without 10 μg/mL bleomycin. WST-8 assay was performed after treatment with bleomycin for the indicated time. Quantification of relative growth rates was performed at 96 h ( n = 4). G HCT116 XRCC4 KO cells stably expressing WT or T308A XRCC4 were all treated with 10 μg/mL bleomycin and were transfected with either control vector or OGA. WST-8 assay was performed after treatment with bleomycin for the indicated time ( n = 5). H HCT116 XRCC4 KO cells stably expressing WT or T308A XRCC4 were treated with or without 10 μg/mL bleomycin. WST-8 assay was performed after treatment with bleomycin for the indicated time ( n = 4). A – H Data were represented as means ± SD. Statistical significance was determined by the two-tailed Student’s t -test ( *P < 0.05, **P < 0.01, ***P < 0.001).

Article Snippet: At each time point following cell attachment, 10 μL of Quanti-Max WST-8 cell viability assay solution (Biomax, #QM1000, Korea) was added to each well, and the cells were maintained in incubation for 1 h at 37 °C.

Techniques: Western Blot, Immunoprecipitation, Quantitative RT-PCR, Stable Transfection, Expressing, Knock-Out, Mutagenesis, Transfection, Control, Plasmid Preparation, Two Tailed Test

Journal: Cell Death & Disease

Article Title: O -GlcNAcylation of XRCC4 controls its stability and confers resistance to DNA double-strand break damage in cancer cells

doi: 10.1038/s41419-025-08209-4

Figure Lengend Snippet: A HCT116 XRCC4 KO cells stably expressing WT or T308A XRCC4 were transfected with OGA. WST-8 assay was performed after growth for the indicated time ( n = 5). B HCT116 XRCC4 KO cells stably expressing WT or T308A XRCC4 were subject to a Transwell invasion assay. Invaded cells were measured by WST-8 assay ( n = 7). C HCT116 XRCC4 KO cells stably expressing WT or T308A XRCC4 were subject to colony formation assays. Colony formation was quantified by using a 485/520 nm filter set ( n = 8). D Each group of BALB/c nude mice ( n = 6 per group) were injected with 1 × 10 7 of the mentioned cells into the hypodermis. Tumor size and weight were quantified 60 days after injection. A – D Data were represented as means ± SD. Statistical significance was determined by the two-tailed Student’s t -test or one-way ANOVA for multiple comparisons ( **P < 0.01, ***P < 0.001).

Article Snippet: At each time point following cell attachment, 10 μL of Quanti-Max WST-8 cell viability assay solution (Biomax, #QM1000, Korea) was added to each well, and the cells were maintained in incubation for 1 h at 37 °C.

Techniques: Stable Transfection, Expressing, Transfection, Transwell Invasion Assay, Injection, Two Tailed Test