Desktop computer-based image analysis of cell surface fluorescence patterning from a photographic source

We report the use of standard computer-based image analysis technology to analyze, from a photographic source, individual cell surface receptor site patterns using fluorochrome labeling. The model used in this study was a Strongylocentrotus purpuratus sea urchin embryo labeled with fluorescein isothiocyanate-conjugated wheat germ agglutinin (FITC-WGA) (0.5 mg/ml for 5 min at 15 degrees C incubated with fertilization membrane free embryos). Image capture was performed using desktop-type digital scanning, and the images were imported into Adobe Photoshop for study. All images analyzed were derived from a single photographic negative: 1) the actual micrograph printed from the negative and scanned into a Macintosh IIx computer; 2) the scanned negative itself; and, 3) a high resolution scanning process used with a Kodak Photo CD. Patterns were analyzed using the densitometry feature of Photoshop, and were similar enough from all 3 scanned images to indicate that any of the 3 scanning processes can be used for fluorescence patterning analysis. Adobe Photoshop was also used to highlight, for closer analysis, the fluorescence patterns by producing 3-D effects, border mapping and transition area detailing. The desktop image analysis procedures described here to study fluorescence patterning require no expensive scientific hardware or software.