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. 2009 Jan 21;29(7):1972–1986. doi: 10.1128/MCB.01590-08

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FIG. 4. — The anti-histone H3.3 antibody immunoprecipitates HA-tagged histone H3.3 but not HA-tagged histone H3. Immunoprecipitation with the anti-histone H3 antibody (ab1791; Abcam) or our anti-histone H3.3 antibody was carried out with sonicated lysates of HEK cells that had been transiently transfected with recombinant HA-tagged H3.3 or H3 (constructs kindly donated by Chunyuan Jin and Gary Felsenfeld). Cells (∼5 × 10⁶) were transiently transfected with FuGENE HD at a ratio of 3:2 (microliters of FuGENE to micrograms of DNA) according to the manufacturer's instructions (Roche Diagnostics). At 72 h posttransfection, cells were collected for immunoprecipitation experiments. As a negative control (−Ab), the immunoprecipitation assay was performed in the absence of either antibody. Samples were then subjected to Western blotting with an anti-HA tag-specific polyclonal antibody (ab9110; Abcam).