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Anti erg

Manufactured by Abcam
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Anti-ERG is a lab equipment product used for the detection and analysis of the ERG (ETS-related gene) protein. ERG is a transcription factor that plays a role in various cellular processes. This product is designed to aid researchers in studying the expression and function of the ERG protein in their research applications.

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Lab products found in correlation

Anti gapdh, by Merck Group (2 mentions) Anti p53 do 1, by Santa Cruz Biotechnology (2 mentions) Iblot gel transfer stacks nitrocellulose, by Thermo Fisher Scientific (1 mentions) Pager gold precast gel, by Lonza (1 mentions) Iblot gel transfer system, by Thermo Fisher Scientific (1 mentions) Anti pten, by Abcam (1 mentions) Anti β actin antibody, by Santa Cruz Biotechnology (1 mentions) Bca protein assay reagent, by Bio-Rad (1 mentions) Hoechst 33342, by Thermo Fisher Scientific (1 mentions) Alexa 488, by Thermo Fisher Scientific (1 mentions) Protein block serum free, by Agilent Technologies (1 mentions) Alexa fluor 488 igg, by Thermo Fisher Scientific (1 mentions) Anti ezh2, by BD (1 mentions) Anti c myc antibody, by Cell Signaling Technology (1 mentions) Puromycin, by Merck Group (1 mentions) Tet system approved fetal bovine serum, by Takara Bio (1 mentions) Anti p21 waf1 cip1, by Cell Signaling Technology (1 mentions) Anti e2f1, by Cell Signaling Technology (1 mentions) Hrp labeled secondary antibody, by Proteintech (1 mentions) Anti claudin 5, by Abcam (1 mentions)

Spelling variants of this product

Rabbit anti-ERG (6 citations) Anti-ERG antibody (4 citations)

10 protocols using anti erg

1

Western Blot Analysis of ERG, PTEN, and β-Actin

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Cells were lysed in modified RIPA buffer (Santa Cruz) supplemented with PMSF, protease inhibitor and sodium orthovanadate. Protein concentration of the lysates were determined using BCA protein assay reagent (Bio-Rad); 30μg of the extracted protein was mixed with Laemmli sample buffer containing β-ME, denatured, separated using 10% PAGEr Gold Precast Gels (Lonza), and transferred using iBlot Gel Transfer system (Invitrogen) and iBlot Gel Transfer Stacks Nitrocellulose (Invitrogen). Anti-ERG (#2805-1, Epitomics), anti-PTEN (#S-0271, Epitomics), anti-β-actin antibody (Santa Cruz Biotech), antibodies were used at 1:1000 dilution with blocking with 5% skim milk. After incubation with primary antibodies overnight at 4°C, horseradish peroxidase–labeled secondary antibodies were then applied to the membranes for 1 h at room temperature. Signals were visualized using ECL Western blotting detection substrate (Thermo Scientific).
Shao L., Wang J., Karatas O.F., Feng S., Zhang Y., Creighton C.J, & Ittmann M. (2018). Fibroblast growth factor receptor signaling plays a key role in transformation induced by the TMPRSS2/ERG fusion gene and decreased PTEN. Oncotarget, 9(18), 14456-14471.
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2

Immunofluorescence Imaging of EZH2 and ERG

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To perform immunofluorescence cells were grown on glass cover-slips, fixed with 4% formaldehyde, permeabilized with 1:1 methanol–acetone, incubated with anti-ERG (Epitomics), anti-EZH2 (BD Biosciences), and anti-mERG as primary antibodies and then with anti-rabbit Alexa 594 or anti-mouse Alexa 488 (Invitrogen). To perform immunofluorescence for anti-pS21 EZH2 (Bethyl Laboratories) cells were grown in μ-slide (chambered coverslip) with 8 wells (ibidi). At a later stage, cells were fixed and permeabilized with methanol–acetone (1:1) solution. After cells blocked for 10 min with blocking buffer (Dako, Protein Block Serum- Free, Ref. X0909) and incubated for 1 h at room temperature with anti-pS21 EZH2 antibody (Bethyl Laboratories). Then incubation with a secondary goat anti-rabbit Alexa Fluor 488 IgG (Invitrogen) followed. Antibodies were diluted in Antibody Diluent (Boster, AR1016). Nuclear visualization was carried out with Hoechst Staining (Hoechst 33342, Thermo Scientific). Images were obtained using the confocal microscope.
Zoma M., Curti L., Shinde D., Albino D., Mitra A., Sgrignani J., Mapelli S.N., Sandrini G., Civenni G., Merulla J., Chiorino G., Kunderfranco P., Cacciatore A., Kokanovic A., Rinaldi A., Cavalli A., Catapano C.V, & Carbone G.M. (2021). EZH2-induced lysine K362 methylation enhances TMPRSS2-ERG oncogenic activity in prostate cancer. Nature Communications, 12, 4147.
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3

Inducible ERG Expression in LNCaP Cells

Cited 2 times
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LNCaP cell line was transduced with an inducible lentiviral ERG construct (LNCaP-lentivirus TMPRESS2:ERG3 inducible) to establish stable doxycycline-inducible ERG expressing LnTE3 cell line [2 (link), 16 (link)]. The cell lines were cultured in RPMI 1640, supplemented with 10% Tet System Approved Fetal Bovine Serum (Clontech Laboratories, Inc. Mountain View, CA, USA) and puromycin (Sigma, St. Louis, MO, USA) with or without doxycycline (Dox, 1 μg/ml) as per requirements and characterized as described [2 (link), 16 (link)]. Antibodies used were as follows: anti-GAPDH (Millipore MAB374), anti-ERG (Abcam ab92513), anti- p21Waf1/Cip1, anti-E2F1 and anti- c-Myc Antibody (Cell Signaling 2946, 3742 and 9402, respectively), anti-p53 DO1 (Santa Cruz biotech, sc126), and anti-NKX3.1 (Biocare Medical SKU 422).
Kumar P., Chakraborty J., Sukumar G., Dalgard C., Chatterjee R, & Biswas R. (2019). Comparative RNA-seq analysis reveals dys-regulation of major canonical pathways in ERG-inducible LNCaP cell progression model of prostate cancer. Oncotarget, 10(42), 4290-4306.
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4

Protein Expression Analysis in Cultured Cells

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Cultured cells (5 × 106) were lysed on ice using PMSF lysate buffer. Samples containing 40–60 μg of protein were subjected to thermal denaturation and SDS-PAGE and were then transferred to PVDF membranes. After the membrane was sealed with a blocking solution (TBST containing 5% skim milk) for 60 min, it was incubated with the primary antibody: anti-ERG (Abcam, England, Ca#133264), anti-VE-cadherin (Abcam, Ca# ab33168), and anti-claudin-5 (Abcam, Ca# ab15106) overnight and then with the HRP-labeled secondary antibody (Proteintech, USA, Ca# SA00001-1) at 37°C or 1 h. Finally, the BeyoECLPlus chemiluminescence reagent (Beyotime, China, P0018) was used to visualize the protein bands.
Zheng B., Wang H., Cui G., Guo Q., Si L., Yan H., Fang D., Jiang L., Jiang Z, & Zhou J. (2020). ERG-Associated lncRNA (ERGAL) Promotes the Stability and Integrity of Vascular Endothelial Barrier During Dengue Viral Infection via Interaction With miR-183-5p. Frontiers in Cellular and Infection Microbiology, 10, 477.
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5

Protein Expression Analysis Protocol

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Protein extraction and immunoblot analysis were performed using the standard protocol. Antibodies used were as follows: anti GAPDH (Millipore MAB374), Anti-ERG (Abcam ab92513), Anti-AR (Active Motif 39781), anti-PMEPA1 (ABNOVA H00056937-M01), Anti-PSA (DakoCytomation A05662), anti-p53 DO1 (Santa Cruz biotech, sc126), anti-SIRT1 and Acetyl-p53(Lys379) (Cell Signaling 2493 and 2570).
Kumar P., Sharad S., Petrovics G., Mohamed A., Dobi A., Sreenath T.L., Srivastava S, & Biswas R. (2016). Loss of miR-449a in ERG-associated prostate cancer promotes the invasive phenotype by inducing SIRT1. Oncotarget, 7(16), 22791-22806.
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6

Antibodies and Reagents for Protein Analysis

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Antibodies against CITED2, PRMT5, β-tubulin, WDR77, RioK1, p300, β-CTN, Vimentin, TWIST, Snail, N-Cad, and ZEB1 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA); anti-NCL, anti-dimethyl-arginine and anti-acetyl-lysine from Upstate Biotechnology (Lake Placid, NY); anti-p-AKT, anti-AKT and anti-E-cadherin from Cell Signaling (Danvers, MA); anti-FLAG, anti-MYC, and anti-HA from Sigma-Aldrich (St. Louis, MO); and anti-ERG and anti-α-SMA from Abcam (Cambridge, MA). MK2206 was purchased from Selleckchem. Human recombinant proteins of CITED2, PRMT5, P300, and NCL were purchased from Origene. Fetal bovine serum (FBS), dithiothreitol, G418 disulfate salts (G418), EPZ015666, Leptomycin B, Bay-11-7082, LY294002, Cycloheximide, Wortmannin, and others were obtained from Sigma-Aldrich. Sources and dilution factors of antibodies used are summarized in Supplementary Table 1.
Shin S.H., Lee G.Y., Lee M., Kang J., Shin H.W., Chun Y.S, & Park J.W. (2018). Aberrant expression of CITED2 promotes prostate cancer metastasis by activating the nucleolin-AKT pathway. Nature Communications, 9, 4113.
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7

Tissue Preparation and Immunohistochemistry

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After euthanasia, mice were subjected to transcardial perfusion with PBS + heparin (Sigma-Aldrich) and 4% paraformaldehyde. Tissue samples were collected and fixed in 4% PFA in 1× PBS overnight and embedded in paraffin. Sections were stained with hematoxylin-eosin for analysis. For immunohistochemistry, antibody detection was performed using the Elite ABC kit and DAB substrate according to the manufacturer's instructions (Vector Laboratories) and counterstained with hematoxylin (Thermo Fisher Scientific). Primary antibodies used were rat anti-FLAG (Biolegend 637319), anti-CD31 (Dianova DIA-310), anti-endomucin (Santa Cruz Biotechnology sc-65495), rabbit anti-S100a6 (Novus Biologicals NBP1-89388), anti-ERG (Abcam ab133264), anti-CD34 (Abcam ab81289), anti-Ki67 (Leica Biosystems NCLKi67p), anti-cytokeratin, wide spectrum screening (Agilent Z0622), anti-CRYAB (Invitrogen PA1-16950), and mouse anti-PDGFA (Santa Cruz Biotechnology sc-9974). Alexa488-, Alexa568-, and Alexa647-conjugated secondary antibodies (Molecular Probes) were used for immunofluorescence.
Driskill J.H., Zheng Y., Wu B.K., Wang L., Cai J., Rakheja D., Dellinger M, & Pan D. (2021). WWTR1(TAZ)-CAMTA1 reprograms endothelial cells to drive epithelioid hemangioendothelioma. Genes & Development, 35(7-8), 495-511.
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8

Western Blot Analysis of Cell Signaling

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Lysates were prepared using RIPA buffer (10 mM Tris-Cl (pH 8.0), 1 mM EDTA, 1% (v/v) Triton X-100, 0.1% (w/v) sodium deoxycholate, 0.1% (w/v) SDS and 140 mM NaCl) supplemented with protease inhibitor tablets (ThermoFisher, UK). Equal protein samples were separated on 10% (v/v) SDS polyacrylamide gel electrophoresis gels and transferred to PVDF membranes (Sigma Aldrich) which were blocked and probed overnight at 4 °C with 1:1000 anti-ERG (Abcam, UK), 1:10,000 anti-β-actin (Abcam), 1:2500 anti-GAPDH (Millipore, UK), 1:1000 β-catenin, 1:1000 anti-c-Myc, 1:1000 anti-cyclin D1 or 1:1000 anti-phospho-LRP6 (all Cell Signalling Technologies, UK) primary antibodies. Membranes were incubated in 1:2000 HRP-linked anti-rabbit or anti-mouse IgG secondary antibody (NEB) for 2 h at room temperature. Membranes were incubated in Luminata Forte Western HRP substrate (Millipore) for chemiluminescent detection prior to image acquisition using a Li-Cor Odyssey imaging system.
Li L., Hobson L., Perry L., Clark B., Heavey S., Haider A., Sridhar A., Shaw G., Kelly J., Freeman A., Wilson I., Whitaker H., Nurmemmedov E., Oltean S., Porazinski S, & Ladomery M. (2020). Targeting the ERG oncogene with splice-switching oligonucleotides as a novel therapeutic strategy in prostate cancer. British Journal of Cancer, 123(6), 1024-1032.
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9

Multimarker Immunohistochemistry for Cardiovascular Tissues

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Anti-CX40 (1:500; Alpha Diagnostics Int. Inc.; Cat: CX40-A), αSMA-FITC (1:300; Sigma; Cat: F3777), anti-ERG (1:500; Abcam; Cat: ab92513), CXCR4 (1:125; BD Pharminogen; Cat: 551852 and 1:300; abcam; Cat: 124824), Anti-PH3 (1:500; Millipore; Cat: 06-570), Anti-Myomesin (1:500; DSHB; B4-C), Anti-WGA (1:100, Invitrogen, Cat: W32466), Anti-ENDOMUCIN (1:300, Invitrogen, Cat: 14-5851-82), Anti-VEGFR2 (1:125, R&D Systems, Cat: AF644). Secondary reagents were Alexa fluor-conjugated antibodies (405, 488, 555, 633) from Life Technologies used at 1:250 dilutions.
Das S., Goldstone A.B., Wang H., Farry J., D’Amato G., Paulsen M.J., Eskandari A., Hironaka C.E., Phansalkar R., Sharma B., Rhee S., Shamskhou E.A., Agalliu D., de Jesus Perez V., Woo Y.J, & Red-Horse K. (2019). A unique collateral artery development program promotes neonatal heart regeneration. Cell, 176(5), 1128-1142.e18.
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10

Prostate Cancer TMA Immunostaining Protocol

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Prostate cancer TMAs were purchased from US Biomax, Inc. As indicated by US Biomax, Inc (https://www.biomax.us/FAQs#q10), all tissues were collected under the highest ethical standards with the donors being formally informed and with their consent obtained. Given that US Biomax, Inc has the patient consent form in place already, the investigators of the current study did not acquire additional informed written consent from the subjects. However, the patient studies were conducted in accordance the ethical guidelines of Declaration of Helsinki and approved by Mayo Clinic Institutional Review Board (IRB). TMA specimens were used for antigen retrieval and immunostaining as described previously (36 (link)). Primary antibodies used were anti-FOXO1 (Bethyl) and anti-ERG (Abcam). Staining intensity and staining percentage for each tissue was graded using a set of criteria. Intensity was graded 0 to 3, with 0 being no staining, 1 low staining, 2 medium staining, and 3 strong staining. A final staining index (SI) score for each staining was obtained by multiplying values obtained from staining percentage and intensity and used for correlation analysis.
Yang Y., Blee A.M., Wang D., An J., Pan Y., Yan Y., Ma T., He Y., Dugdale J., Hou X., Zhang J., Weroha S.J., Zhu W.G., Wang Y.A., DePinho R.A., Xu W, & Huang H. (2017). Loss of FOXO1 cooperates with TMPRSS2-ERG overexpression to promote prostate tumorigenesis and cell invasion. Cancer research, 77(23), 6524-6537.
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