FIGURE 5.

Data were acquired from a cell expressing mRFP1 for a period of 5.5 min. The data were divided into 10 consecutive segments, each 0.5 min long, and histogrammed. Each of the 10 histograms was fit to a single-species model to obtain the brightness in each channel (A) and the number of molecules (B) as a function of the data segment number (or data acquisition time). The brightness of mRFP1 is larger than that shown in Fig. 3 B due to the use of a higher excitation power. At this excitation power, some photobleaching of mRFP1 occurs. Thus we see a reduction in the number of molecules in consecutive histograms. However, the brightness is unaffected by the photobleaching and remains constant across the data acquisition time.