EpiCypher是一家为表观遗传学和染色质生物学研究提供高质量试剂和工具的专业制造商。EpiCypher推出的CUT&RUN级别的HA Antibody符合EpiCypher的“CUTANA Compatible”标准，可用于核酸酶靶向切割和释放（CUT&RUN）和/或靶向剪切及转座酶技术（CUT&Tag）的基因组定位。HA抗体可用于利用HA标记的靶蛋白研究。HA-Tag抗体在表达3xHA-标记的GATA3转录因子的乳腺癌细胞中产生与GATA3 DNA结合motif（Fig.2）重叠的CUT&RUN峰（Fig.1）。

产品详情

反应种属: HA Epitope (YPYDVPDYA)

宿主来源: Rabbit

实验应用: CUT&RUN, WB

免疫原: A synthetic HA peptide (sequence: YPYDVPDYA)

克隆性: Polyclonal

保存温度: Stable for 1 year at 4℃ from date of receipt

运输温度: DO NOT FREEZE!! Frozen cold packs.

产品形式: Antigen affinity-purified antibody in phosphate buffered saline (PBS), 0.09% sodium azide

数据示例

Figure 1: HA Tag peaks in CUT&RUN

CUT&RUN was performed as described above. Peaks were called using MACS2. (A) Heatmaps show GATA3-3xHA peaks relative to IgG and H3K4me3 control antibodies in aligned rows ranked by intensity (top to bottom) and colored such that red indicates high localized enrichment and blue denotes background signal. (B) The number of GATA3-3xHA peaks that fall into distinct classes of functionally annotated genomic regions are shown.

Figure 2: HA-tagged transcription factor binding motif analysis in CUT&RUN

(A) Homer analysis determined that the GATA3 consensus motif, represented as a sequence logo position weight matrix, was enriched under GATA3-3xHA CUT&RUN peaks. (B) The number of GATA3-3xHA peaks containing GATA3 consensus motifs from panel A is represented by a Venn Diagram. (C-D) Two representative loci show overlap of GATA3-3xHA peaks with the consensus motifs noted by tick marks beneath the tracks.

Figure 3: Western blot data

E. coli cells expressing a multi-tag fusion protein were used to prepare whole cell lysates. The indicated amounts (ng) of lysate were loaded onto a 4-20% SDS-PAGE gel and analyzed under standard western blot conditions using HA Tag antibody at a dilution of 1:25,000.

Figure 4: Target-specific epitope cleavage of HA Tag antibody in CUT&RUN was determined using DNA-barcoded recombinant nucleosome spike-in controls

(A) A panel of recombinant nucleosomes was created where various epitope tags (3xTY1, 3xFLAG, 3xHA) were fused to the histone H3 tail. The fused nucleosomes and an unmodified control were immobilized to streptavidin beads (SA Bead) and spiked into CUT&RUN samples alongside ConA bead immobilized MDA-MB-231 cells expressing GATA3-3xHA (Figure 1). HA Tag antibody and pAG-MNase (EpiCypher 15-1016) were then added to release antibody-bound nucleosomes into solution through pAG-MNase mediated cleavage of the linker DNA (light blue). This approach provided a defined experimental control to assess whether the HA Tag antibody selectively cleaved the target epitope with high specificity and minimal background. (B) CUT&RUN sequence reads were aligned to the unique DNA 'barcodes' corresponding to each nucleosome in the spike-in panel. Data are expressed as the percent of reads recovered relative to the intended target (3xHA, set to 100%). This analysis confirms that the HA Tag antibody specifically liberated the target epitope-tagged nucleosome into solution.

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