Anti-Histone H3 (mono methyl K36) antibody - ChIP Grade

• ICC/IF

Lab

Immunocytochemistry/ Immunofluorescence - Anti-Histone H3 (mono methyl K36) antibody - ChIP Grade (AB9048)

ab9048 staining Histone H3 (mono methyl K36) in HeLa cells. The cells were fixed with 100% methanol (5 min), permeabilized with 0.1% PBS-Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1%PBS-Tween for 1h. The cells were then incubated overnight at 4°C with ab9048 at 0.1µg/ml and ab7291, Mouse monoclonal [DM1A] to alpha Tubulin - Loading Control. Cells were then incubated with ab150081, Goat polyclonal Secondary Antibody to Rabbit IgG - H&L (Alexa Fluor® 488), pre-adsorbed at 1/1000 dilution (shown in green) and ab150120, Goat polyclonal Secondary Antibody to Mouse IgG - H&L (Alexa Fluor® 594), pre-adsorbed at 1/1000 dilution (shown in pseudocolour magenta). Nuclear DNA was labelled with DAPI (shown in blue). Also suitable in cells fixed with 4% paraformaldehyde (10 min).Image was acquired with a high-content analyser (Operetta CLS, Perkin Elmer) and a maximum intensity projection of confocal sections is shown.

• ChIP

Lab

ChIP - Anti-Histone H3 (mono methyl K36) antibody - ChIP Grade (AB9048)

Chromatin was prepared from HeLa cells according to the Abcam X-ChIP protocol. Cells were fixed with formaldehyde for 10 minutes. The ChIP was performed with 25μg of chromatin, 2μg of ab9048 (blue), and 20μl of Protein A/G sepharose beads. No antibody was added to the beads control (yellow). The immunoprecipitated DNA was quantified by real time PCR (Taqman approach). Primers and probes are located in the first kb of the transcribed region.

• WB

AbReview25029****

Western blot - Anti-Histone H3 (mono methyl K36) antibody - ChIP Grade (AB9048)

All lanes:

Western blot - Anti-Histone H3 (mono methyl K36) antibody - ChIP Grade (ab9048) at 1/1400 dilution

All lanes:

Whole cell lysate prepared from Drosophila BG3 cells at 500000 Cells

Secondary

All lanes:

HRP donkey anti-rabbit monoclonal at 1/20000 dilution

Predicted band size: 15 kDa

Observed band size: 17 kDa,42 kDa

true

Exposure time: 30s

Image courtesy of an anonymous Abreview.

• WB

Unknown

Western blot - Anti-Histone H3 (mono methyl K36) antibody - ChIP Grade (AB9048)

Secondary ab : Alexa Fluor 680 Goat anti-rabbit IgG

0.5μg histone prep used per lane

Primary antibody :
Lane 1 : ab9048 (Histone H3 Mono Methyl K36) 1/500
Lane 2 : ab9048 (Histone H3 Mono Methyl K36) 1/500 + ab1783 (ab9048) (Histone H3 Mono Methyl K36) peptide 1μg/ml
Lane 3 : ab9048 (Histone H3 Mono Methyl K36) 1/500 + ab1794 (ab9049) (Histone H3 Di Methyl K36) peptide 1μg/ml
Lane 4 : ab9048 (Histone H3 Mono Methyl K36) 1/500 + ab1785 (ab9050) (Histone H3 Tri Methyl K36) peptide 1μg/ml
Lane 5 : ab9048 (Histone H3 Mono Methyl K36) 1/500 + ab2623 (Histone H3 (23-34) – unmodified) peptide 1μg/ml
Lane 6 : ab9048 (Histone H3 Mono Methyl K36) 1/500 + ab1340 (ab8895) (Histone H3 Mono methyl K4) peptide 1μg/ml

ab9048 specifically recognise

All lanes:

Western blot - Anti-Histone H3 (mono methyl K36) antibody - ChIP Grade (ab9048) at 1/500 dilution

Lane 1:

Histone prep at 0.5 µg

Lane 2:

Histone prep at 0.5 µg with Human Histone H3 (mono methyl K36) peptide (<a href='/en-us/products/unavailable/human-histone-h3-mono-methyl-k36-peptide-ab1783'>ab1783</a>)

Lane 3:

Histone prep at 0.5 µg with Human Histone H3 (di methyl K36) peptide (<a href='/en-us/products/proteins-peptides/human-histone-h3-di-methyl-k36-peptide-ab1784'>ab1784</a>)

Lane 4:

Histone prep at 0.5 µg with Human Histone H3 (tri methyl K36) peptide (<a href='/en-us/products/proteins-peptides/human-histone-h3-tri-methyl-k36-peptide-ab1785'>ab1785</a>)

Lane 5:

Histone prep at 0.5 µg with Human Histone H3 (unmodified ) peptide (<a href='/en-us/products/proteins-peptides/human-histone-h3-unmodified-peptide-ab2623'>ab2623</a>)

Lane 6:

Histone prep at 0.5 µg with Human Histone H3 (mono methyl K4) peptide (<a href='/en-us/products/proteins-peptides/human-histone-h3-mono-methyl-k4-peptide-ab1340'>ab1340</a>)

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab6721'>ab6721</a>) at 1/5000 dilution

Predicted band size: 15 kDa

false