Anti–AML1–ETO polyclonal antibody

Catalog #: 25208
Size: 100 µl
$365
Host Species: Rabbit
Clonality: polyclonal
Background:
The AML1 (RUNX1) (UniProtKB/Swiss–Prot entry Q01196) – ETO (RUNX1T1) (UniProtKB/Swiss–Prot entry Q06455) fusion protein arises due to a translocation between chromosome 8 and 22 (t(8;21)(q22;q22)). This translocation is one of the most frequent karyotypic abnormalities observed in acute myeloid leukaemia. It produces a chimerical gene made up of the 5’–region of AML1and the 3’–region of ETO. The chimerical protein is thought to associate with the nuclear corepressor/histone deacetylase complex to block hematopoietic differentiation.
Description:
Polyclonal antibody raised in rabbit against the AML1–ETO fusion protein using a KLH–conjugated synthetic peptide.
Synonym(s):
RUNX1–RUNX1T1 fusion protein
Assay Conditions:

ChIP results obtained with the antibody directed against AML1-ETO
ChIP assays were performed using Kasumi-1 cells, the antibody against AML1-ETO (Cat. # 25208) and optimized primer pairs for qPCR. Sheared chromatin from 1.25 million cells and 4 µl of antibody were used per ChIP experiment. QPCR was performed using primers specific for the FUT7, NFE2 and OGG1 genes. Figure 1 shows the occupancy, calculated as the ratio + control/background for which the promoter of the H2B gene was used.

ChIP-seq results obtained with the antibody directed against AML1-ETO
ChIP was performed as described above. The immunoprecipitated DNA of 6 ChIP assays was pooled and analyzed with an Illumina Genome Analyzer. Library preparation, cluster generation and sequencing were performed according to the manufacturer's instructions. The 32 bp tags were aligned to the human reference genome (hg18) using the ELAND algorithm. Figure 2 shows the results of the complete chromosome 3 and three genomic regions surrounding the OGG1, FUT7 and NFE2 genes, respectively. The position of the PCR amplicon is indicated with an arrow.

Determination of the antibody titer
To determine the titer of the antibody, an ELISA was performed using a serial dilution of the antibody directed against AML1-ETO (Cat. # 25208). The plates were coated with the peptides used for immunization of the rabbit. By plotting the absorbance against the antibody dilution (Figure 3), the titer of the antibody was estimated to be 1:32,750.

Western blot analysis using the antibody directed against AML1-ETO
Nuclear extracts of SKNO-1 cells (15 µg) were analyzed by Western blot using the antibody against AML1-ETO (Cat. # 25208) diluted 1:1,000 in TBS-Tween containing 5% skimmed milk. The marker (in kDa) is shown on the left, the position of the protein of interest is indicated on the right. 

Concentration:
100 µl
Formulation:
Whole antiserum from rabbit containing 0.05% azide
Species Reactivity:
Human
Purification:
Whole antiserum
Immunogen:
synthetic peptide
Format:
aqueous solution
Storage / Stability:

Store at –80°C for up to 2 years. Centrifuge after first thaw to maximize product recovery. Aliquot to avoid repeated freeze/thaw cycles. Aliquots may be stored at –20°C for at least one month.

Application(s):
ChIP/ChIP – seq (4 µl/ChIP)
ELISA (1:500)
WB (1:1000)
Notes:
The optimal antibody amount per IP should be determined by the end–user. We recommend testing 1–5 µg per IP
Warning(s):
Avoid freeze/thaw cycles
Scientific Category:
Antibodies
Product Type:
Antibody
Data shown is lot-specific. Contact us for specific information on other lots.

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