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> H3K27me3 (Ser28) Polyclonal Antibody

H3K27me3 (Ser28) Polyclonal Antibody

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Referencia bs-53053R

embalaje : 100uL

Marca : Bioss


ChIP assays were performed using HeLa cells treated with colcemid, the H3K27me3S28p antibody (bs-53053R) and optimized PCR primer pairs for qPCR. ChIP was performed using sheared chromatin from 10,000 cells. A titration of the antibody consisting of 1, 5 and 10 \u03bcl per ChIP experiment was analysed. Additionally, ChIP was performed after incubation of the antibody with 5 nmol blocking peptide (cat. No. sp-091-050) for 1 hour at room temperature. IgG (5 \u03bcg\/IP) was used as negative IP control. QPCR was performed with primers for the promoters of the active genes c-fos and RPL30 and for the inactive gene MYOD. The figure shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).
ChIP assays were performed using HeLa cells treated with colcemid, the H3K27me3S28p antibody (bs-53053R) and optimized PCR primer pairs for qPCR. ChIP was performed using sheared chromatin from 10,000 cells. A titration of the antibody consisting of 1, 5 and 10 \u03bcl per ChIP experiment was analysed. Additionally, ChIP was performed after incubation of the antibody with 5 nmol blocking peptide (cat. No. sp-091-050) for 1 hour at room temperature. IgG (5 \u03bcg\/IP) was used as negative IP control. QPCR was performed with primers for the promoters of the active genes c-fos and RPL30 and for the inactive gene MYOD. The figure shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).
To determine the titer, an ELISA was performed using a serial dilution of the H3K27me3S28p antibody (bs-53053R). The antigen used was a peptide containing the histone modifications of interest. By plotting the absorbance against the antibody dilution, the titer of the antibody was estimated to be 1:8,300.
To determine the titer, an ELISA was performed using a serial dilution of the H3K27me3S28p antibody (bs-53053R). The antigen used was a peptide containing the histone modifications of interest. By plotting the absorbance against the antibody dilution, the titer of the antibody was estimated to be 1:8,300.
A Dot Blot analysis was performed to test the cross reactivity of the H3K27me3S28p antibody (bs-53053R) with peptides containing other modifications of histone H3 and H4 and unmodified sequences from histone H3. One hundred to 0.2 pmol of the peptides were spotted on a membrane. The antibody was used at a dilution of 1:20,000. The figure shows a high specificity of the antibody for the peptide containing the modifications of interest.
A Dot Blot analysis was performed to test the cross reactivity of the H3K27me3S28p antibody (bs-53053R) with peptides containing other modifications of histone H3 and H4 and unmodified sequences from histone H3. One hundred to 0.2 pmol of the peptides were spotted on a membrane. The antibody was used at a dilution of 1:20,000. The figure shows a high specificity of the antibody for the peptide containing the modifications of interest.
Hela cells were stained with the H3K27me3S28p antibody (bs-53053R) and with DAPI. Cells were fixed with formaldehyde, permeabilized with sodium citrate and Triton X100 and blocked with PBS containing 2.5% BSA. Left side: cells were immunofluorescently labelled with the H3K27me3S28p antibody (diluted 1:200 and incubated for 1 hour at room temperature) followed by goat anti-rabbit antibody conjugated to DyLight 488. Right side: the nuclei were stained with DAPI, which specifically labels DNA. Phosphorylation of H3 on serine 28 is increased during late G2 phase and reaches a maximum in metaphase cells. This may explain the different staining intensities of different cells.
Hela cells were stained with the H3K27me3S28p antibody (bs-53053R) and with DAPI. Cells were fixed with formaldehyde, permeabilized with sodium citrate and Triton X100 and blocked with PBS containing 2.5% BSA. Left side: cells were immunofluorescently labelled with the H3K27me3S28p antibody (diluted 1:200 and incubated for 1 hour at room temperature) followed by goat anti-rabbit antibody conjugated to DyLight 488. Right side: the nuclei were stained with DAPI, which specifically labels DNA. Phosphorylation of H3 on serine 28 is increased during late G2 phase and reaches a maximum in metaphase cells. This may explain the different staining intensities of different cells.
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