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Biotechniques. 1993 Jan;14(1):42-8.

Three-color immunofluorescence imaging of Drosophila embryos by laser scanning confocal microscopy.

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  • 1Howard Hughes Medical Institute, Laboratory of Molecular Biology, Madison, WI 53706.


We present a simple means for triple-labeling biological specimens by immunofluorescence using a laser scanning confocal microscope for imaging with a krypton/argon laser as a light source. Three separate images of fluorescein-, lissamine rhodamine- and cyanine-5-labeled antibodies are collected and subsequently merged to form the triple-labeled image, which is displayed at full-image resolution (24 bit) on a second image processing system. The technique is illustrated using immunofluorescence localization of three segmentation proteins in Drosophila embryos.

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