Quantification of protein mobility and associated reshuffling of cytoplasm during chemical fixation.

摘要: To understand cellular functionalities, it is essential to unravel spatio-temporal patterns of molecular distributions and interactions within living cells. The technological progress in fluorescence microscopy now allows principle measure these with sufficient spatial resolution. However, high resolution imaging comes long acquisition times phototoxicity. Therefore, physiological live cell often unfeasible chemical fixation employed. Yet, methods have not been rigorously investigated, terms pattern preservation, at the which cells can be imaged. A key parameter for this time required until complete. During time, are under unphysiological conditions decay. We demonstrate here that formaldehyde takes more than one hour cytosolic proteins cultured Other small aldehydes, glyoxal acrolein, did perform better. Associated this, we found a distinct displacement lipids, including their loss from Fixations using glutaraldehyde were faster four minutes retained most cytoplasmic proteins. Surprisingly, autofluorescence produced by was almost completely absent supplementary addition without compromising speed. These findings indicate, processes actually reliably imaged after certain fixation.