IMMUNOFLUORESCENT VISUALIZATION OF THE EXCRETORY AND GUT SYSTEM OF SCHISTOSOMA-MANSONI BY CONFOCAL LASER-SCANNING MICROSCOPY

Conventional epifluorescence microscopy (CEM) and confocal laser scanning microscopy (CLSM) were used to visualize the excretory system and the gut on whole organisms of different life-cycle stages of Schistosoma mansoni. To visualize the gut system, an anti-circulating anodic antigen (CAA) monoclonal antibody (MAb) (120-1B10-A) was used, whereas the excretory system was immunohistochemically stained with an antiflame cell MAb (51-4H8-A) and with a recently described anti-egg MAb (114-5B1-A). The CEM procedure resulted in clear images at low magnification but the signal-to-noise ratio on the higher magnification images was very poor. Using CLSM on the adult worm, the 114-581-A MAb demonstrated a well-defined system of canals that could be morphologically identified as the excretory system. The flame eels terminating the branches of the excretory canals showed a clear immunoreactivity with the 114-5B1-A MAb as well as with the specific flame cell MAb. The gut system could be visualized, using an anti-CAA MAb, as two well-defined bands throughout the length of the parasite. Application of the 114-5B1-A MAb on cercariae revealed a strong fluorescence on the cercarial surface, whereas no immunoreactivity could be detected on internal structures. Whole eggs showed a bright fluorescence of the egg shell, whereas miracidia showed immunoreactivity of the germinal cells located in the center of the organism. The CLSM procedure, especially with the recently introduced fast photon-counting option, provides a superior tool to investigate the three-dimensional localization of different epitopes on immunofluorescently stained whole mounts of multicellular organisms in comparison with CEM. The advent of new visualization techniques to exactly localize epitopes of newly discovered MAbs can be of great significance in the development of new screening tests. The identification of epitopes on the different life-cycle stages of this clinically important trematode is valuable for the improvement of therapeutic and prophylactic strategies.