Patterns of Mcm6 and PCNA expression. Molt4 cells were permeabilized with Triton X-100, then fixed with MeOH prior to staining for Mcm6, PCNA, and DNA content. (A) G1 and G2/M clusters are defined by negative PCNA staining at 2C DNA and 4C DNA content, respectively. Positive PCNA staining delimits the S phase compartment. A few 4C G1 cells can be observed. (B) Mcm6 staining is bimodal in G1 phase (arrows), and the off-loading of Mcm proteins from chromatin during S phase is apparent. Note: the immunofluorescence axes are plotted on a hyperlog scale that produces a linear to log gradient and accommodates negative numbers and zero . (C) a bivariate histogram of PCNA versus Mcm6 expression divides the G1/S interface into distinct clusters with very few cells classified as intermediate. (D) The pattern of Mcm6 expression in Molt4 cells varied with growth rate. In rapidly dividing cells, the high Mcm6 population was more pronounced (arrow) than in more slowly growing cells (Figure 1C, late G1). To simplify the bivariate patterns, G2/M cells were not included in C and D.