Glutaraldehyde (GA) fixation of TFSMs. HeLa cells were incubated with BODIPY-FL-C 12 -sphingomyelin (BODIPY-FL-C 12 -SM), TFSM 1 or TFSM 2 for 2 h, bSMase-treated, detached, fixed, and permeabilized with TX-100 (as indicated). Fixation was done either with paraformaldehyde (PFA) or PFA and GA. TFSM then were BODIPY-FL-DBCO-stained and analyzed by flow cytometry. n = 3 (BODIPY-FL-C 12 -SM, TFSM 1 PFA and TFSM 2 PFA), n = 6 ( TFSM 1 GA + PFA, TFSM 2 GA + PFA). Statistics: Two-way ANOVA and Sidaks ( b ) or Tukeys ( e ) multiple comparisons. Two-sided unpaired Students t test ( c ). Bars represent means SD. n corresponds to biological replicates. Source data and detailed statistics are provided as a Source Data file.
