H3K18ac Polyclonal Antibody

$HeLa cells were stained with the Bioss antibody against H3K18ac (cat. No. bs-53137R) and with DAPI. Cells were fixed with 4% formaldehyde for 10\u2019 and blocked with PBS\/TX-100 containing 5% normal goat serum and 1% BSA. The cells were immunofluorescently labeled with the H3K18ac antibody (left) diluted 1:200 in blocking solution followed by an anti-rabbit antibody conjugated to Alexa488. The middle panel shows staining of the nuclei with DAPI. A merge of the two stainings is shown on the right.$

HeLa cells were stained with the Bioss antibody against H3K18ac (cat. No. bs-53137R) and with DAPI. Cells were fixed with 4% formaldehyde for 10\u2019 and blocked with PBS\/TX-100 containing 5% normal goat serum and 1% BSA. The cells were immunofluorescently labeled with the H3K18ac antibody (left) diluted 1:200 in blocking solution followed by an anti-rabbit antibody conjugated to Alexa488. The middle panel shows staining of the nuclei with DAPI. A merge of the two stainings is shown on the right.

$Western blot was performed on whole cell (25 \u03bcg, lane 1) and histone extracts (15 \u03bcg, lane 2) from HeLa cells, and on 1 \u03bcg of recombinant histone H2A, H2B, H3 and H4 (lane 3, 4, 5 and 6, respectively) using the Bioss antibody against H3K18ac (cat. No. bs-53137R). The antibody was diluted 1:500 in TBS-Tween containing 5% skimmed milk.$

Western blot was performed on whole cell (25 \u03bcg, lane 1) and histone extracts (15 \u03bcg, lane 2) from HeLa cells, and on 1 \u03bcg of recombinant histone H2A, H2B, H3 and H4 (lane 3, 4, 5 and 6, respectively) using the Bioss antibody against H3K18ac (cat. No. bs-53137R). The antibody was diluted 1:500 in TBS-Tween containing 5% skimmed milk.

To test the cross-reactivity of the Bioss antibody against H3K18ac (cat. No. bs-53137R), a Dot Blot analysis was performed with peptides containing other histone modifications and the unmodified H3K18. One hundred to 0.2 pmol of the respective peptides were spotted on a membrane. The antibody was used at a dilution of 1:5,000. The figure shows a high specificity of the antibody for the modification of interest.

To determine the titer of the antibody, an ELISA was performed using a serial dilution of the Bioss antibody against H3K18ac (cat. No. bs-53137R). The antigen used was a peptide containing the histone modification of interest. By plotting the absorbance against the antibody dilution, the titer of the antibody was estimated to be 1:4,300.

$ChIP was performed as described above using 1 \u03bcg of H3K18ac Polyclonal Antibody (cat. No. bs-53137R). The IP\u2019d DNA was subsequently analyzed on an Illumina Genome Analyzer. Library preparation, cluster generation, and sequencing were performed according to the manufacturer\u2019s instructions. The 36 bp tags were aligned to the human genome using the ELAND algorithm. The figure shows the peak distribution along the complete human X-chromosome and a zoom-in to a 600 kb region (figure A and B), and in two regions on chromosome 14 and 3 surrounding the c-fos and EIF4A2 positive control genes (figure C and D).$

ChIP was performed as described above using 1 \u03bcg of H3K18ac Polyclonal Antibody (cat. No. bs-53137R). The IP\u2019d DNA was subsequently analyzed on an Illumina Genome Analyzer. Library preparation, cluster generation, and sequencing were performed according to the manufacturer\u2019s instructions. The 36 bp tags were aligned to the human genome using the ELAND algorithm. The figure shows the peak distribution along the complete human X-chromosome and a zoom-in to a 600 kb region (figure A and B), and in two regions on chromosome 14 and 3 surrounding the c-fos and EIF4A2 positive control genes (figure C and D).

$ChIP assays were performed using HeLa cells, treated with TSA, the Bioss antibody against H3K18ac (cat. No. bs-53137R) and optimized PCR primer pairs for qPCR. ChIP was performed with a ChIP-seq kit, using sheared chromatin from 1,000,000 cells. A titration consisting of 1, 2, 5 and 10 \u03bcg of antibody per ChIP experiment was analyzed. IgG (2 \u03bcg\/IP) was used as a negative IP control. Quantitative PCR was performed with primers for the promoters of the active EIF4A2 and c-fos genes (positive controls), and for the inactive MYOD1 gene and the Sat2 satellite repeat (negative controls). The figure shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).$

ChIP assays were performed using HeLa cells, treated with TSA, the Bioss antibody against H3K18ac (cat. No. bs-53137R) and optimized PCR primer pairs for qPCR. ChIP was performed with a ChIP-seq kit, using sheared chromatin from 1,000,000 cells. A titration consisting of 1, 2, 5 and 10 \u03bcg of antibody per ChIP experiment was analyzed. IgG (2 \u03bcg\/IP) was used as a negative IP control. Quantitative PCR was performed with primers for the promoters of the active EIF4A2 and c-fos genes (positive controls), and for the inactive MYOD1 gene and the Sat2 satellite repeat (negative controls). The figure shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).

$ChIP assays were performed using HeLa cells treated with TSA, \r\nH3K18ac Polyclonal Antibody (cat. No. bs-53137R) and optimized PCR primer pairs for qPCR. ChIP was performed with a ChIP-seq kit using sheared chromatin from 1,000,000 cells. A titration consisting of 1, 2, 5 and 10 \u03bcg of antibody per ChIP experiment was analyzed. IgG (2 \u03bcg\/IP) was used as a negative IP control. Quantitative PCR was performed with primers for the promoters of the active EIF4A2 and c-fos genes, used as positive controls, and inactive MYOD1 gene and the Sat2 satellite repeat, used as negative controls. The figure shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).$

ChIP assays were performed using HeLa cells treated with TSA, \r\nH3K18ac Polyclonal Antibody (cat. No. bs-53137R) and optimized PCR primer pairs for qPCR. ChIP was performed with a ChIP-seq kit using sheared chromatin from 1,000,000 cells. A titration consisting of 1, 2, 5 and 10 \u03bcg of antibody per ChIP experiment was analyzed. IgG (2 \u03bcg\/IP) was used as a negative IP control. Quantitative PCR was performed with primers for the promoters of the active EIF4A2 and c-fos genes, used as positive controls, and inactive MYOD1 gene and the Sat2 satellite repeat, used as negative controls. The figure shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).

Applications

WB
ELISA
IF
dot-blot
ChIP-seq

Reactivity

Human
Mouse

Overview
Catalog #	bs-53137R
Product Name	H3K18ac Polyclonal Antibody
Applications	WB, ELISA, IF, dot-blot, ChIP-seq
Reactivity	Human, Mouse
Specifications
Conjugation	Unconjugated
Host	Rabbit
Source	Polyclonal antibody raised in rabbit against the region of histone H3 containing the acetylated lysine 18 (H3K18ac), using a KLH-conjugated synthetic peptide
Clonality	Polyclonal
Isotype	Not Determined
Concentration	0.8ug/ul
Purification	Affinity purified polyclonal antibody in PBS containing 0.05% azide and 0.05% ProClin 300.
Storage Buffer	PBS with 0.05% sodium azide and 0.05% Proclin300.
Storage Condition	Store at -20°C for 12 months.
Target
Gene ID	8350
Swiss Prot	P68431
Background	Core component of nucleosome. Nucleosomes wrap and compact DNA into chromatin, limiting DNA accessibility to the cellular machineries which require DNA as a template. Histones thereby play a central role in transcription regulation, DNA repair, DNA replication and chromosomal stability. DNA accessibility is regulated via a complex set of post-translational modifications of histones, also called histone code, and nucleosome remodeling.
Application Dilution
WB	1:300-5000
ELISA	1:500-1000
IF
dot-blot
ChIP-seq