Fluorescence surface contour plot

Figure 12.5 Tri-dimensional fluorescence maps (a) surface contour plot and (b) projected surface plot, highlighting the iso-intensity contour lines.

The third problem that needs to be controlled is that of spectral cross-over and the possibility of incorrect instrument compensation. As an example of a case in which controls for nonspecific staining, autofluorescence, and compensation are all critical, let us look at the staining of B lymphocytes for the CD5 marker present with only low density on their surface. As well as the problems created by nonspecific staining and by autofluorescence, the problem of spectral cross-over between fluorescein and phycoerythrin can particularly confuse the interpretation of results from this kind of experiment. Look at Figure 6.5. What we are interested in is the number of B lymphocytes that possess the CD5 surface antigen. These cells will appear in quadrant 2 of a contour plot of fluorescein fluorescence... 

Figure 7.2. The variability in EEM fluorescence for whole DOM collected from surface waters compared to ultrafiltered permeate (<1 kDa MWCO) and retentate (>1 kDa MWCO) fractions. (A-C) Susquehanna River water collected at the head of the Chesapeake Bay. (D-E) Coastal seawater collected from the Middle Atlantic Bight. The boxes and letters on each EEM contour plot refer to the fluorescent peak region designations from Table 7.1.

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