Optimization of cell transfer and fixation for immunofluorescence analysis using spiked samples.

<p>(A) Representative images to show a CellTracker Green pre-labeled PC3 cell with damaged morphology (green arrow) after Cytospin (lower panel), compared with a round pre-labeled PC3 cell (green arrow) in direct cell loading method (upper panel). (B) and (C) Using both (B) PBS-4% paraformaldehyde and (C) KCl-acetone cell transfer and fixation methods, CK expression (green signals) on PC3 cells (green arrows) and CD45 expression (red signals) on lymphocytes (red arrows) were clearly detected with strong and specific fluorescence signals. (D) Percentage of cells retained after immunofluorescence staining process for cells with different re-suspension solutions and fixations. Both 70% and pure acetone fixation in combination with KCl re-suspension of cells achieved a retention rate of more than 90%. (E) Harvest rates of spiked PC3 cells using different cell transfer and fixation methods. KCl-acetone had the highest rate, Cytospin had a lower but more stable rate and PBS-4% paraformaldehyde had the lowest and most unstable rate.