Atto 488 goat anti mouse igg

The following primary rabbit antibodies were purchased from Abcam: anti-H1.2, ab17677; H1.5, ab18208; HMGB2, ab124670; Ki67, ab15580; H3S10p, ab5176; CTCF, ab128873; SMC2, ab10412; RAD21, ab154769; H1x, ab31972. From Cell Signaling: anti-HMGN1, #5692; HMGN2, #9437. From Sigma-Aldrich: anti-H1.4, H7665. From Millipore: anti-H2A, #07-146. The following mouse antibodies were purchased from Abcam: anti-H3, ab24834; H4, ab31830; H2B, ab52584. From Active Motif: anti-H3, #61475. mAb PL2-6 was a gift from M. Monestier (Temple Univ.), see previous use [26,27]. Secondary antibodies included Invitrogen Alexa 568 goat anti-rabbit IgG, Sigma-Aldrich Atto 488 goat anti-rabbit IgG and Atto 488 goat anti-mouse IgG.

Cells were seeded onto coverslips, incubated for 20 h before drug treatment or viral transduction, and then fixed in PTEMF buffer containing 20 mM PIPES, 4% paraformaldehyde, 0.2% Triton-X, 10 mM EGTA and 1 mM MgCl₂ for 15 min. Primary antibodies included anti-HSP70 (GeneTex, Hsinchu, Taiwan or StressMarq, Victoria, Canada), anti-α-tubulin (GTX112141, GeneTex or T5168, Sigma, St. Louis, MO, USA), anti-γ-tubulin (T6557 or T3559, Sigma), anti-EB1 (E3406, Sigma), anti-pericentrin (ab4448 or ab28144, Abcam, Cambridge, UK), anti-CEP215 (IHC-00063, Bethyl, Cambridge, UK), anti-CEP164 (HPA037606, Sigma), anti-CEP152 (ab183911, Abcam), anti-PLK1 (No. 33-1700, Invitrogen), and anti-phospho-T210-PLK1 (PA1-126, ThermoFisher, Waltham, MA, USA). Alexa-Fluor 488-, 568-, 633-, or 647-conjugated goat anti-mouse or anti-rabbit IgG were purchased from Invitrogen, and Atto-488 goat anti-mouse IgG was from Sigma. Samples were mounted in Fluoromount-G from SouthernBiotech (Birmingham, AL, USA) for confocal or structured illumination microscopy (SIM) imaging. For ground state depletion (GSD) imaging, samples were mounted in DPBS (Sigma) containing 5% glucose, 100 mM 2-mercaptoethylamine (Sigma), 0.8 mg/ml glucose oxidase and 40 μg/ml catalase, and sealed with Twinsil (Picodent, #13001000, Wipperfürth, Germany).