Human serum albumin fraction 5

Human serum albumin (Fraction V) and glutaraldehyde 8% aqueous solution were purchased from Sigma (Steinheim, Germany). Magnetite particles were friendly provided by Merck KGaA (Darmstadt, Germany). All reagents were of analytical grade and used as received.

Cells were grown on 13 mm coverslips in 12-well plates, washed in DPBS (14190136, Gibco™) before fixation with 4% paraformaldehyde and storage in DPBS. Before immunocytochemical staining, cells were washed in DPBS, and blocked using ADST [antibody diluent solution—triton: DPBS, 0.1 M L-Lysine (303341000, Thermo Scientific™), 1% w/v Human Serum Albumin Fraction V (12668-10GM, Sigma-Aldrich), Triton X-100 (A16046.AP, Thermo Scientific Alfa Aesar)] and 5% human serum (H3667, Sigma Aldrich) for 30 min. Cells were washed and primary antibodies at 2.5 µg/ml (suspended in ADST with 2% human serum) were applied overnight. After washing, secondary antibodies at 5 µg/ml and 4′,6-diamidino-2-phenylindole (DAPI, D1306, Invitrogen™) at 1 µg/ml (suspended in ADST with 2% human serum) were applied for 1 h, before mounting coverslips using Dako mounting medium (S302380-2, Agilent). Antibodies were sourced from Abcam: Rb anti-Ki67 (ab15580, ab16667), Ms anti-γH2AX (ab26350), Alexa Fluor ® 555 Goat pAb to Rb (ab150078, ab150086) and Alexa Fluor ® 488 Goat pAb to Ms (ab150117). Images were captured using the Leica DM4 B Upright Microsope at 10 × magnification and were counted manually using Leica Application Suite X 2019 3.7.1.21655v software (Leica Microsystems, Wetzlar, Germany).