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. 2014 May 1;10(5):e1004347. doi: 10.1371/journal.pgen.1004347

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© 2014 Ruckenstuhl et al

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.

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Fluorescent microscopy of acidic vacuoles during chronological aging of MET+, Δmet2, and Δmet2/Δatg5 strains, by means of quinacrine accumulation and statistical analysis thereof. (>1000 cells of each strain from 3 to 5 independent samples at each time point were evaluated. Only cells with acidic vacuoles without an additionally stained cytoplasm were counted as positive, resulting in cell counts that represent cells which have a clearly intact pH-homeostasis. Positively counted cells are indicated by white arrowheads) (A and B). (C) Statistical analysis of fluorescent microscopy of acidic vacuoles by means of quinacrine accumulation. Strains Δmet2 and Δmet2/Δatg5 were grown to stationary phase under excess of methionine and shifted to media with the indicated amounts of methionine (>500 cells from each strain from 2 independent samples) and assayed for quinacrine accumulation after ∼20 hours (D) Chronological aging of the MET⁺ strain overexpressing Vma1p or Vph2p. Cell death was measured via propidium iodide staining of cells that have lost integrity and subsequent flow cytometry analysis (BD LSRFortessa) (n = 6 to 8). See also Figure S6.