Anti-Histone H2A.Z antibody - ChIP Grade

• WB

Ap

Western blot - Anti-Histone H2A.Z antibody - ChIP Grade (AB4174)

All lanes:

Western blot - Anti-Histone H2A.Z antibody - ChIP Grade (ab4174) at 1 µg/mL

Lane 1:

Calf Thymus Histone Preparation Nuclear Lysate at 0.5 µg

Lane 2:

HeLa (Human epithelial carcinoma cell line) Whole Cell Lysate at 10 µg

Lane 3:

NIH/3T3 (Mouse embryonic fibroblast cell line) Whole Cell Lysate at 10 µg

Lane 4:

PC-12 (Rat adrenal pheochromocytoma cell line) Whole Cell Lysate at 10 µg

Secondary

All lanes:

Goat polyclonal to Rabbit IgG - H&L - Pre-Adsorbed (HRP) (ab65484) at 1/3000 dilution

Predicted band size: 14 kDa

Observed band size: 16 kDa,36 kDa

true

Exposure time: 3min

• ChIP

Unknown

ChIP - Anti-Histone H2A.Z antibody - ChIP Grade (AB4174)

Chromatin was prepared from HeLa (Human epithelial cell line from cervix adenocarcinoma) cells according to the Abcam X-ChIP protocol.

Cells were fixed with formaldehyde for 10 min. The ChIP was performed with 25 μg of chromatin, 2 μg of ab4174 (blue), and 20 μl 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 for active and inactive loci, Sybr green approach for heterochromatic loci). Primers and probes are located in the first kb of the transcribed region.

• ChIP

PubMed

ChIP - Anti-Histone H2A.Z antibody - ChIP Grade (AB4174)

Changes in nucleosome occupancy upon LPS induction at a putative distal enhancer and promoter of IL1A, kinetics of nucleosome removal, and changes in histone modifications.

Chromation from mouse bone marrow derived macrophages. ChIP experiments with antibodies against H3 (dark blue bars), H2A.Z (light blue, (ab4174 at 4μg), H3K4me1 (green, ab8895 at 1 μg), H3K4me3 (yellow, ab8580 at 1 μg) and H3K27ac (red, ab4729 at 1 μg). For these experiments cross-linked chromatin was lightly digested with MNase before incubation with the respective antibodies to increase resolution of the ChIP signal and the data was normalized to a region in the ORF of RPL4. Changes upon LPS induction in histone binding and histone modifications at the enhancers and promoters of IL12B and IL1A as well as at a control region in the GAPDH pseudo gene are shown as fold over levels found before induction. For H3K27ac the changes 1.5 h after LPS induction, and for all other histone variants and modifications the changes after 3 h of induction are shown. The error bars show the SEM of at least 3 independent experiments. Statistical significance of the changes in H3K4me3 and H3K27ac upon LPS induction compared to levels found prior to induction determined by Student's T-tests is indicated (*P<0.05; **P<0.01).

Gjidoda et al PLoS One. 2014 Apr 4;9(4):e93971. doi: 10.1371/journal.pone.0093971. eCollection 2014. Fig 2. Reproduced under the Creative Commons license http://creativecommons.org/licenses/by/4.0/

• WB

Unknown

Western blot - Anti-Histone H2A.Z antibody - ChIP Grade (AB4174)

All lanes:

Western blot - Anti-Histone H2A.Z antibody - ChIP Grade (ab4174) at 1/500 dilution

All lanes:

Calf thymus histone lysate

Secondary

All lanes:

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

Predicted band size: 14 kDa

Observed band size: 15 kDa

false

• WB

AbReview47162****

Western blot - Anti-Histone H2A.Z antibody - ChIP Grade (AB4174)

Blocked with 3% BSA for 1 hour at 20°C.

Primary incubation in TBS tween + 3% BSA at 20°C for 1 hour.

All lanes:

Western blot - Anti-Histone H2A.Z antibody - ChIP Grade (ab4174) at 1/1000 dilution

Lane 1:

Native recombinant octamers K562 (Human chronic myelogenous leukemia cell line from bone marrow) cells at 3 µg

Lane 2:

Native recombinant octamers K562 cells at 1.5 µg

Lane 3:

Recombinant Human octamers containing H2A at 1 µg

Lane 4:

Recombinant Human octamers containing H2A at 0.5 µg

Lane 5:

Recombinant Human octamers containing H2A.Z.2.1 at 0.5 µg

Lane 6:

Recombinant Human octamers containing H2A.Z.1 at 0.5 µg

Secondary

All lanes:

HRP-conjugated donkey anti-rabbit IgG polyclonal at 1/10000 dilution

Predicted band size: 14 kDa

Observed band size: 15 kDa

true

Exposure time: 5min

This image is courtesy of an Abreview submitted by Ragnhild Eskeland