Ab81278

Manufactured by Abcam

Sourced in China

Ab81278 is a monoclonal antibody that recognizes human Bax protein. Bax is a pro-apoptotic member of the Bcl-2 family of proteins and plays a crucial role in the mitochondrial apoptosis pathway.

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Lab products found in correlation

6 protocols using ab81278

Western Blot Analysis of Key Cellular Proteins

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Total protein was extracted from cultured cells using RIPA buffer. The primary polyclonal antibody against P14 (ab3648, Abcam), P15 (ab53034, Abcam), P16 (ab81278, Abcam), CBX7 (ab21873, Abcam), GAPDH (60004-1, ProteinTech), or green fluorescent protein (GFP) (ProteinTech, 50430-2-AP; China) was applied at 1:5,000 to 1:1,000 dilutions. The signals were visualized using the Enhanced Chemiluminescence Kit (Millipore) and Alpha Imager system.

Li Z., Qiao J., Ma W., Zhou J., Gu L., Deng D, & Zhang B. (2022). P14AS upregulates gene expression in the CDKN2A/2B locus through competitive binding to PcG protein CBX7. Frontiers in Cell and Developmental Biology, 10, 993525.

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Immunostaining for Cell Proliferation and Lineage Markers

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The cells were fixed with 4% paraformaldehyde for 10 min, permeabilized with 0.1% PBS-TritonX-100 for 5 min, and then blocked with 1% BSA/10% normal goat serum/0.3 M glycine in 0.1% PBS-Tween for 1 h. The cells were then incubated overnight at 4°C with anti-Ki67 antibody (ab16667, Abcam, 1 : 200)/anti-SOX10 antibody (GTX35085, GeneTex, 1 : 100) or anti-P16 antibody (ab81278, Abcam, 1 : 200)/anti-SOX10 antibody, followed by further incubation at 20–30°C for 1 h with goat anti-rabbit IgG H&L (FITC, ab6717, Abcam, 1 : 1000) and goat anti-mouse IgG H&L (Cy3 ®) preadsorbed (ab97035, Abcam, 1 : 500) secondary antibodies. Nuclear DNA was labeled blue with DAPI.

Wei B., Gu J., Duan R., Gao B., Wu M., Zhou S., Huang X, & Xie F. (2021). Transcriptome Analysis of Large to Giant Congenital Melanocytic Nevus Reveals Cell Cycle Arrest and Immune Evasion: Identifying Potential Targets for Treatment. Journal of Immunology Research, 2021, 8512200.

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Xenograft Protein Expression Analysis

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Freshly frozen xenograft tissue samples were lysed in radioimmunoprecipitation assay buffer using a TissueLyzer II (Qiagen) homogenizer at 4°C following the manufacturer’s instructions. SDS–polyacrylamide gel electrophoresis was performed by standard methods, and polyvinylidene difluoride membranes were blocked with bovine serum albumin (BSA) 2.5% and PBS Tween 0.5% for 60 min at room temperature and probed overnight at 4°C with the following antibodies: PAR (1:4000; 4336-BPC-100, Trevigen), glyceraldehyde-3-phosphate dehydrogenase (GAPDH; 1:40,000; MAB374, Millipore), MDR1 (1:200; sc-13131, Santa Cruz Biotechnology), MGMT (1:1000; 2739, Cell Signaling Technology), SLFN11 (1:500; sc-374339, Santa Cruz Biotechnology), p57 (1:500; ab75974, Abcam), p16 (1:2000; ab81278, Abcam) and p21 (1:1000; 2947, Cell Signaling Technology), CYCLIN A2 (1:1000; 4656, Cell Signaling Technology), CYCLIN B1 (1:1000; 12231, Cell Signaling Technology), H3pSer10 (1:1000; 06-570, Millipore), and MYC (1:500, sc-40, Santa Cruz Biotechnology). Membranes were imaged with a Syngene G:BOX, band densitometry was performed using FIJI, and ratio to loading control (GAPDH) was calculated.

Stanzione M., Zhong J., Wong E., LaSalle T.J., Wise J.F., Simoneau A., Myers D.T., Phat S., Sade-Feldman M., Lawrence M.S., Hadden M.K., Zou L., Farago A.F., Dyson N.J, & Drapkin B.J. (2022). Translesion DNA synthesis mediates acquired resistance to olaparib plus temozolomide in small cell lung cancer. Science Advances, 8(19), eabn1229.

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Protein Expression Analysis in Cells

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Twenty micrograms of total protein was used per sample. The following primary antibodies were used: γH2AX (1:1,000, 2577S; Cell Signaling Technology, Inc., Danvers, MA, USA); CDKN2A/p16^ink4a (1:1,000, ab 81278; Abcam, Cambridge, UK); SIRT1 (1:1,000, ab 32441; Abcam); SIRT6 (1:1,000, ABE102; EMD Millipore, Billerica, MA, USA); anti-perilipin A (1:1,000, P1998-200UL; Sigma-Aldrich Co., St Louis, MO, USA); anti-MyoD1 (1:1,000; Abcam); cleaved caspase-3 (Asp175; 1:1,000, 9661S; Cell Signaling Technology, Inc.). GAPDH was used as a load control (1:5,000; Santa Cruz Biotechnology Inc., Dallas, TX, USA). The optical densities were quantified using a densitometer and Image-Pro Plus.

Lakhdar R., McGuinness D., Drost E.M., Shiels P.G., Bastos R., MacNee W, & Rabinovich R.A. (2018). Role of accelerated aging in limb muscle wasting of patients with COPD. International Journal of Chronic Obstructive Pulmonary Disease, 13, 1987-1998.

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Western Blot Analysis of Cell Lysates

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Approximately 1 × 10 6 cells were collected and lysed to obtain protein lysates. After blocking with 5% fat-free milk overnight at 4°C, the membrane was incubated with primary antibodies (anti-P16, ab81278, Abcam; anti-TERT, ab32020, Abcam; anti-GAPDH, #60004-1, Protein Tech Group, China) for one hour at room temperature, and then incubated with the appropriate goat anti-rabbit (A0208, Beyotime, Beijing, China) or goat anti-mouse (A0216, Beyotime) secondary antibodies at room temperature for one hour.

Zhang X., Li P., Gan Y., Xiang S., Gu L., Zhou J., Zhou X., Wu P., Zhang B, & Deng D. (2024). Driving effect of P16 methylation on telomerase reverse transcriptase-mediated immortalization and transformation of normal human fibroblasts. Chinese medical journal.

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Protein Extraction and Western Blotting

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Experimental cells were harvested by using the total protein extraction buffer (Tris 7.58 g/L, SDS 20 g/L, β-mercaptoethanol 20 mL/L, Glycerol 100 mL/L, Bromophenol Blue 0.2 g/L, pH6.8) and the lysates were denatured immediately at 100°C for 10 min. Subsequently, equal amounts of protein extracts were subjected to separation by SDS-PAGE, and subsequent visualization was performed by Western blotting with indicated antibodies. The monoclonal antibody of NFE2L1 (D5B10) was purchased from CST, USA. The antibody of Ubiquitin (ab134953), WNT6 (ab154144), CDKN1A (ab109520) and CDKN2A (ab81278) were purchased from Abcam, USA. The antibody of FZD8 (D261206), WNT9A (D121400), CDC25A (D155203), β-catenin (D260137) were purchased from Sangon Biotech, China. The antibody of CCND1 (GB11079) was purchased from Servicebio, China. The β-actin (GB11001, Servicebio, China) and Tubulin (ab210797, Abcam, USA) were served as internal control to correct the mass of samples.

Qiu L., Yang Q., Li P., Xiaowen Z., Yang Y., Zhou X., Gou S., Wenjie Z., Li G., Ren Y., Zhao W., Wu Y., Qi Y., & Gao Y. (2020). Function expansion of antitumor transcriptional activator NFE2L1 by the original discovery of its non-transcription factor activity.

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