Influence of different fixation reagents on NIH 3T3 fibroblasts morphology: An AFM study
B.Codan, V.Martinelli, S.Maggiolino, L.Mestroni, O.Sbaizero
The Atomic force microscope (AFM) has recently become an attractive tool for biological analysis, especiallyfor the characterization of bothmorphological and mechanical properties. In this study, we investigatedthe influence of different fixation reagentsonNIH 3T3 fibroblast cells’ morphology. Three protocolswere selected: paraformaldehyde, methanol and acetone. Height, perimeter, area, volume and roughness of cells were measured using an AFM. As expected, different fixation protocols produce differentmorphological changes in cells. However, to preserve fibroblasts’ morphology, the methods applying aldehyde fixations performed much better than those using either methanol or acetone. Based on quantitative assessments, fibroblasts fixed with paraformaldehyde showed the closest volume, area and perimeter values to living cells. Roughness data and a Western blot test confirmed that methanol fixation produces larger cell membrane damage, when compared to the other two fixatives.
AFM; Cellmorphology; Fixatives.