Histone H3K4me3 Antibody, SNAP-ChIP Certified *DISCONTINUED*

Applications Key: ChIP-Chromatin IP; E-ELISA; FACS-Flow cytometry; IF-Immunofluorescence; IHC-Immunohistochemistry; IP-Immunoprecipitation; WB-Western Blotting

Reactivity Key: B-Bovine; Ce-C. elegans; Ch-Chicken; Dm- Drosophila; Eu-Eukaryote; H-Human; M-Mouse; Ma-Mammal; R-Rat; Sc-S.cerevesiae; Sp-S. pombe; WR-Wide Range (predicted); X-Xenopus; Z-Zebrafish

Representative SNAP-ChIP-seq results: Cumulative histogram plot and heatmap of signal intensity depict H3K4me3 ChIP-seq data aligned to annotated transcription start sites (TSS, +/- 3.0 kb; left). Two representative genomic regions depicting H3K4me3 peak structure and distribution are shown (right). Data shown are representative of H3K4me3 ChIP antibody (EpiCypher Catalog No. 13-0028) and are not lot-specific. Native ChIP-seq was performed using K562 cells as described (Shah et al., Mol Cell 2018) with SNAP-ChIP^TM K-MetStat Spike-in (Catalog No. 19-1001) nucleosome controls added prior to chromatin digestion to confirm antibody specificity and ChIP efficiency. Paired-end sequencing libraries were prepared using the NEBNext^® Ultra^TM II DNA Library Prep Kit for Illumina^®. ChIP libraries were sequenced on an Illumina^® NextSeq. Sequencing reads were aligned to the human genome using Bowtie 2 (Johns Hopkins University). Bigwig files of read enrichment in binned genomic regions (signal intensity) flanking the indicated gene features were used to create a cumulative histogram plot and heatmap of signal intensity ( www.basepairtech.com). Gene browser shots were generated using the Integrative Genomics Viewer (IGV, Broad Institute) with the window size denoted (top).
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SNAP-ChIP qPCR Data: Histone H3K4me3 antibody (3 μg) was tested in a native ChIP experiment with chromatin from HEK-293 cells (~1x10⁶ cells) with the SNAP-ChIP K-MetStat Panel (EpiCypher Catalog No. 19-1001). spiked-in prior to micrococcal nuclease digestion. Specificity (left Y-axis) was determined by qPCR for the DNA barcodes corresponding to modified nucleosomes in the SNAP-ChIP panel (x-axis). Black bar represents antibody efficiency (right y-axis; log scale) and indicates percentage of the target immunoprecipitated relative to input. Error bars represent mean ± SEM in replicate ChIP experiments.
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SNAP-ChIP-seq Data: Histone H3K4me3 antibody (5 μg) was tested in a native ChIP experiment using chromatin from K-562 cells (~1x10⁶ cells) with the SNAP-ChIP K-MetStat Panel (~1x10⁶ cells) with the SNAP-ChIP K-MetStat Panel (EpiCypher Catalog No. 19-1001) spiked-in prior to micrococcal nuclease digestion. Ten nanograms ChIP DNA was subjected to library preparation using the NEBNext® Ultra^TM II DNA Library Prep Kit for Illumina®. ChIP libraries were analyzed by 2x150bp paired end sequencing on an Illumina HiSeq 4000. Paired reads were aligned to the SNAP-ChIP barcodes using the alignment algorithm available at https://www.basepairtech.com/. Specificity was determined by normalizing the read counts for each barcoded nucleosome IP to the corresponding Input and expressing the data as a percent of the nucleosome containing the target PTM (% Target). Error bars represent mean ± SEM for the duplicate DNA barcodes in a single ChIP experiment.
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ELISA Data: Designer Nucleosomes (dNucs) containing the indicated H3K4 modification were used in an ELISA assay with Histone H3K4me3 Antibody (10 μg/mL).
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