For western blotting, protein samples were separated on polyacrylamide gels, blotted to nitrocellulose membranes and probed with the indicated antibodies. The primary antibodies used were mouse anti-H3K9me2 (Abcam ab1220), rat anti-α-tubulin (Sanbio), mouse anti-Flag M2 (Sigma), and rat anti-GFP (ChromoTek). The secondary antibodies used were IRdye680 or IRdye800 conjugated goat anti-rat or goat anti-mouse antibodies (LI-COR). All primary antibodies were diluted 1:1000, and secondary antibodies were diluted 1:10000. Western blots were imaged on an Odyssey CLX imaging system (LI-COR).
Mouse anti h3k9me2
Manufactured by Abcam
Sourced in Belgium
Mouse anti-H3K9me2 is a primary antibody that recognizes the di-methylated form of histone H3 at lysine 9 residue. It is used for the detection and quantification of this histone modification in various research applications.
Automatically generated - may contain errors
Lab products found in correlation
Odyssey clx imaging system, by LI COR (1 mentions)
Mouse anti flag m2, by Merck Group (1 mentions)
Rat anti gfp, by Proteintech (1 mentions)
Irdye 680, by LI COR (1 mentions)
Ab14714, by Abcam (1 mentions)
Ab24684, by Abcam (1 mentions)
Polymer refine detection system, by Leica (1 mentions)
Bond 3 stainer, by Leica (1 mentions)
Mouse anti hif 1α, by Novus Biologicals (1 mentions)
Nb100 132, by Abcam (1 mentions)
Rabbit anti 5hmc, by Active Motif (1 mentions)
Mouse anti 5mc, by Eurogentec (1 mentions)
Bx51 epifluorescence microscope, by Olympus (1 mentions)
Mouse anti gfp, by Roche (1 mentions)
Mouse anti flag, by Merck Group (1 mentions)
Chicken anti gfp, by Abcam (1 mentions)
Mouse anti ha, by BioLegend (1 mentions)
Rabbit anti h3k27me3, by Merck Group (1 mentions)
Rabbit anti h3k9me1, by Abcam (1 mentions)
Goat anti snap25, by Santa Cruz Biotechnology (1 mentions)
5 protocols using mouse anti h3k9me2
1
Comprehensive Western Blot Methodology
2
Immunohistochemical Analysis of Hypoxia Markers
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The staining procedure was performed using a Leica Bond III Stainer (Leica,). Slides were subjected exposed to 10 mM sodium citrate buffer, pH 6.0 for 20 min at 37°C. Slides were incubated with the appropriate primary antibody for 15 min, followed by Polymer Refine Detection System (Leica) processing, including hydrogen peroxidase block, secondary antibody polymer, 3,3’ diaminobenzidine and hematoxylin stain. Specimens were then rinsed for 5 min in tap water. Slides were dehydrated in increasing concentrations of ethyl alcohol and xylene prior to permanent coverslipping in xylene-based media. Primary antibodies were as follows: mouse anti-Hif1α (1:400, Novus Biological), mouse anti-HIF2α (Novus Biologicals, NB100-132, rabbit polyclonal, 1:700) mouse anti-H3K9me2 (1:750, Abcam), rabbit anti-H3K27me2 (ab24684, Abcam, 1:500), rabbit anti-5hmC (39769, Active Motif, 1:4000), and mouse anti-SDHB (ab14714, Abcam, 1:1000).
3
Immunostaining of Epigenetic Modifications
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Embryo samples were fixed in 4% paraformaldehyde [60 (link)] for 1 h at room temperature and permeabilized with 0.5% Triton X-100 in phosphate-buffered saline (PBS) for 30 min. To analyze 5hmC and 5mC, the samples were exposed to 2 M HCl for 15 min at room temperature and then neutralized in 0.5 M Tris-HCl (pH 8.5) for 10 min. Samples were blocked with 1% BSA and 0.2% Triton X-100 in PBS for 2 h and then incubated with primary antibodies overnight at 4 °C, followed by incubation with secondary antibodies for 1 h at room temperature. The nuclei were stained with 4,6-diamidino-2-phenylindole dihydrochloride (DAPI). Images were captured using an Olympus BX51 epifluorescence microscope (Olympus Corp, Tokyo, Japan). The primary antibodies for immunostaining included rabbit anti-5hmC (Active Motif, Carlsbad, CA, USA), mouse anti-5mC (Eurogentec, Liege, Belgium), mouse anti-H3K9me2 (Abcam, Cambridge, UK), and rabbit anti-Dppa3 (Santa Cruz Biotechnology, Santa Cruz, CA, USA).
4
Immunohistochemistry for C. elegans
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The following primary antibodies were used: chicken anti-HTP-3 (1:500) [51 (link)], chicken anti-GFP (1:2,000; Abcam), mouse anti-GFP (1:500; Roche), rabbit anti-GFP (1:500) [52 (link)], mouse anti-HA (1:1,000; Covance/Biolegend), guinea pig anti-SYP-1 (1:200) [53 (link)], mouse anti-H3K9me2 (1:500; Abcam), rabbit anti-SMC-1 (1:200) [54 (link)], rabbit anti-COH-3/4 (1:5,000) [32 (link)], rabbit anti-REC-8 (1:10,000; Novus Biologicals), rabbit anti-HIM-3 (1:200) [14 (link)], rabbit anti-HTP-1/2 (1:500) [16 (link)], and mouse anti-FLAG (1:200; Sigma).
Secondary antibodies conjugated to Alexa dyes 405, 488, 555, or 647, obtained from Molecular Probes, were used at 1:500 dilution (Alexa 488 and 555), 1:200 (Alexa 647), and 1:100 (Alexa 405). In cases in which antibodies raised in mouse and guinea pig were used on the same sample, we used highly cross-absorbed goat anti-mouse secondary antibodies, obtained from Biotium (conjugated to CF488 or CF555, respectively) for secondary detection of the mouse primary antibody in order to avoid cross-reaction against antibodies raised in guinea pig.
Secondary antibodies conjugated to Alexa dyes 405, 488, 555, or 647, obtained from Molecular Probes, were used at 1:500 dilution (Alexa 488 and 555), 1:200 (Alexa 647), and 1:100 (Alexa 405). In cases in which antibodies raised in mouse and guinea pig were used on the same sample, we used highly cross-absorbed goat anti-mouse secondary antibodies, obtained from Biotium (conjugated to CF488 or CF555, respectively) for secondary detection of the mouse primary antibody in order to avoid cross-reaction against antibodies raised in guinea pig.
5
Western Blot and ChIP Antibody Characterization
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The antibodies for Western blot analysis used were: mouse anti-α-tubulin (1:10,000; Sigma, clone B512), rat anti-α-synuclein (1:100; clone 15G7), mouse anti-α-synuclein (1:1000; BD, Synuclein-1), rabbit anti-GFP (1:1000; Santa Cruz), rabbit anti-H3 (1:16000; abcam), rabbit anti-H3K4me3 (1:16000; Abcam), rabbit anti-H3K9me1 (1:16000; Abcam), rabbit anti-H3K9me2 (1:4000; CST), rabbit anti-H3K9me3 (1:16000; Abcam), rabbit anti-H3K27me3 (1:8000; Millipore), rabbit anti-H3K9ac (1:8000; Millipore), rabbit anti-H3K14ac (1:8000; EpiGentek), rabbit anti-H4 (1:16000; Abcam), rabbit anti-H4K12Ac (1:16000; Abcam), rabbit anti-HP1α (1:1000; CST), rabbit anti-EHMT2 (1:2000; abcam), rabbit anti-p38 (1:1000; CST), goat anti-REST (1:250; Santa Cruz) and goat anti-SNAP25 (1:1000; Santa Cruz). The HRP-conjugated secondary antibodies were from Jackson ImmunoResearch Laboratories (1:10,000). The antibodies for chromatin immunoprecipitation (ChIP) used were: rabbit anti-EHMT2 (10 μg; Millipore), mouse anti-H3K9me2 (4 μg; Abcam), and rabbit anti-REST (10 μg; Millipore).
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