Anti top2a

Manufactured by Santa Cruz Biotechnology

Sourced in United States

Anti-TOP2A is a laboratory reagent that targets the DNA topoisomerase 2-alpha (TOP2A) protein. TOP2A is an enzyme involved in the regulation of DNA topology during various cellular processes. Anti-TOP2A is a tool used to study the expression, localization, and function of the TOP2A protein in biological samples.

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5 protocols using anti top2a

Topoisomerase-Dependent Genomic Interactions

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For ChIP-seq we have used anti-Rpb1-NTD (Cell Signaling, D8L4Y), anti-TOP2A (Abcam, ab52934). For ICE, anti-TOP2B (Proteintech, 20549-1-AP), anti-TOP2A (Santa Cruz, SC-365916), anti-TOP1 (Abcam, ab3825). For ICE-IP, anti-TOP2A (abcam, ab52934), anti-TOP1 antibody (abcam, ab109374). For western blot analysis, anti-TOP2B (Proteintech, 20549-1-AP), anti-TOP2A (Santa Cruz, SC-365916), anti-p-p38 (Cell Signaling, 9211) and anti-tubulin (Sigma, T9026), and as secondary antibodies IRDye 680-labeled anti-mouse (LI-COR Biosciences, 926-68070) and IRDye 800-labeled anti-rabbit (LI-COR BIOSCIENCES, 926-32211).
For immunofluorescence, anti-γH2AX (Millipore, 05-636) and anti-H3S10p (Millipore, 06-570). For PLA, anti-NELF-E (Santa Cruz, sc32912), anti-TOP2A (Santa Cruz, SC-365916), and as secondary antibodies Alexa Fluor 488 anti-mouse (Jackson, 715-545-150) and Alexa Fluor 594 anti-rabbit (Jackson, 111-585-003).

Herrero-Ruiz A., Martínez-García P.M., Terrón-Bautista J., Millán-Zambrano G., Lieberman J.A., Jimeno-González S, & Cortés-Ledesma F. (2021). Topoisomerase IIα represses transcription by enforcing promoter-proximal pausing. Cell Reports, 35(2), 108977.

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Chromatin Immunoprecipitation and Western Blot Protocols

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For ChIP, anti-BAF155, anti-BRG1 (J1B), anti-RNA Polymerase II (Santa Cruz N-20), anti-V5 (Life Technologies R960-25), anti-OCT4 (Santa Cruz N-19), and anti-SOX2 (Santa Cruz Y-17) antibodies were used. For etoposide-ChIP, anti-TOP2A (Santa Cruz F-12) antibodies were used. For western blots, anti-BRG1 (J1B), anti-HSP90 (BD Biosciences 68), anti-BAF53a (Novus Biologicals NB100-61628), anti-SS18 (Santa Cruz H-80), anti-V5 (Life Technologies R960-25), and anti-GAPDH (Santa Cruz 6C5) antibodies were used.

Miller E.L., Hargreaves D.C., Kadoch C., Chang C.Y., Calarco J.P., Hodges C., Buenrostro J.D., Cui K., Greenleaf W.J., Zhao K, & Crabtree G.R. (2017). TOP2 synergizes with BAF chromatin remodeling for both resolution and formation of facultative heterochromatin. Nature structural & molecular biology, 24(4), 344-352.

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Protein Expression Analysis in Glioblastoma Cells

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Cell lysates in U-87 MG or T98G cells were extracted, and 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was used to separate total-protein. The nuclear and cytoplasmic extract was prepared using a NE-PER™ Nuclear and Cytoplasmic Extraction Reagent (Thermo Fisher Scientific). After being transferred onto polyvinylidene fluoride (PVDF) membranes, bands were detected after incubation with a primary antibody overnight at 4°C followed by horseradish peroxidase (HRP)-conjugated anti-rabbit secondary antibody for 2 h. All blots were visualized using an enhanced chemiluminescence (ECL) kit (Millipore). Anti-TOP2A, anti-β-catenin, anti-GAPDH, and secondary antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

Liu Y., Ma J., Song J.S., Zhou H.Y., Li J.H., Luo C., Geng X, & Zhao H.X. (2022). DNA topoisomerase II alpha promotes the metastatic characteristics of glioma cells by transcriptionally activating β-catenin. Bioengineered, 13(2), 2207-2216.

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Immunohistochemical Evaluation of Biomarkers

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The primary antibodies used in IHC for TMA slides were as follows: anti-FOXM1 (Proteintech, #13147-1-AP, 1:100 dilution), anti-TOP2A (Santa Cruz, #sc365916, 1:50 dilution), anti-NUF2 (Bioss, #bs-7714 R, 1:100 dilution), anti-KIF18B (Novus, #NBP1-90882, 1:50 dilution). TMAs were stained according to the conventional streptavidin-peroxidase method of IHC (Zymed Laboratories Inc, San Francisco, CA, USA). Two certified pathologists who were blind to the information of patients independently analyzed the IHC results. At high (200×) magnification, the percentage of positive cells was scored as: 1 (0–25%), 2 (26–50%), 3 (51–75%), 4 (>75%), the intensity of positive staining was classified into 4 scales: 0 (negative), 1 (weak), 2 (moderate), 3 (strong). Then, comprehensive score (staining percentage × intensity) was used to evaluate the expression level of signature genes. Disagreements between two pathologists were resolved by consensus. Comprehensive score = 6 was set as cutoff value to define high and low expression group of signature genes.

Yin X., Wang Z., Wang J., Xu Y., Kong W, & Zhang J. (2021). Development of a novel gene signature to predict prognosis and response to PD-1 blockade in clear cell renal cell carcinoma. Oncoimmunology, 10(1), 1933332.

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Chromatin Immunoprecipitation and Western Blot Protocols

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