Application of a digital image-processing tool to the standard SABG assay. A. SABG staining of FSE cells (PD 34) at pH 6.0. B. Image analysis of the field shown in Figure 1A using the Matlab tool described in "Materials". The cell membrane borders and the background control area (small circles) were marked manually. The logarithmic ratio of the cytoplasmic staining intensity (CSI) divided by the background staining intensity (BSI), yields the BGAV. C. Shapiro-Wilk W (SWW) normality test applied to the CSI/BSI values (green line) and to the BGAVs (blue line) obtained from FSE cells at progressively increasing PDs. For any given PD of the FSE cells, the SWW normality value of the logarithmic ratio (BGAV) was higher than the CSI/BSI ratio, indicating a better fit with a normal distribution when applying logarithmic transformation to the CSI/BSI ratio. D. BGAV variation over time. Measurement of BGAVs of FSE cells at different time periods after completion of staining yielded a parabolic distribution of the BGAVs, peaking at 16 hours, and reaching a plateau after 48 hours which remains as such for as long as 2 months following the completion of the staining reaction. This analysis was carried out for FSE cells at two different PDs: PD 43 (green line) and PD 53 (blue line). Hour (h), month (m).