Immunofluorescence staining of Hela cells (treated with 30mM sodium crotonylate for 4h) with CSB-PA010429OA08crHU at 1:7.5, counter-stained with DAPI. The cells were fixed in 4% formaldehyde, permeabilized using 0.2% Triton X-100 and blocked in 10% normal Goat Serum. The cells were then incubated with the antibody overnight at 4°C. The secondary antibody was Alexa Fluor 488-congugated AffiniPure Goat Anti-Rabbit IgG(H+L).

Immunocytochemistry analysis of CSB-PA010429OA08crHU diluted at 1:15 and staining in Hela cells (treated with 30mM sodium crotonylate for 4h) performed on a Leica BondTM system. The cells were fixed in 4% formaldehyde, permeabilized using 0.2% Triton X-100 and blocked with 10% normal goat serum 30min at RT. Then primary antibody (1% BSA) was incubated at 4°C overnight. The primary is detected by a biotinylated secondary antibody and visualized using an HRP conjugated SP system.

Chromatin Immunoprecipitation Hela (4*10⁶, treated with 30mM sodium crotonylate for 4h) were treated with Micrococcal Nuclease, sonicated, and immunoprecipitated with 5µg anti-HIST1H4A (CSB-PA010429OA08crHU) or a control normal rabbit IgG. The resulting ChIP DNA was quantified using real-time PCR with primers against the β-Globin promoter.