New details indicated by different stainings during conjugation of ciliated protozoa Paramecium.

During conjugation of Paramecium caudatum, nuclear events occur in a scheduled program. Morphological studies on nuclear behavior during conjugation of P. caudatum have been performed since the end of the 19th century. Here we report on new details concerning the conjugation of P. caudatum through the staining of conjugating cells with protargol, carbol fuchsin solution, Hoechst 33342 and immunofluorescence labeling with monoclonal antibody of anti-α tubulin. 1) The crescent nucleus is a characteristic of the meiotic prophase of P. caudatum, has an unstained area. We stained this area with protargol, which was separated from the chromatin area and was not detected by the other stainings. 2) In regards to the four meiotic products, it has long been considered that only one product enters the paroral cone region (PC) and survives after meiosis. However, our protargol and immunofluorescence labeling results indicated that PC entrance of the meiotic product happened before the completion of meiosis instead of after. 3) In our previous study, protargol staining indicated the presence of a swollen structure around the central part of the "U" and "V" shaped spindles connecting the two types of prospective pronuclei. However, immunofluorescence labeling with anti-α tubulin antibodies gave a different image from protargol. All these observations form the basis for further studies of their molecular mechanisms.

Localization of stationary pronuclei during conjugation of Paramecium as indicated by immunofluorescence staining.

After the third prezygotic division during conjugation of Paramecium caudatum, migratory and stationary pronuclei are produced. The migratory pronuclei remain in the paroral region tightly against the conjugating boundaries; while the stationary pronuclei are located beside the migratory pronuclei. To date, however, it is not clear what causes this close side-by-side localization between migratory and stationary pronuclei. In the current study, immunofluorescence staining with monoclonal antibody of anti-α tubulin indicated that ''U'' or ''V'' shaped spindles connected the migratory and stationary pronuclei during the third prezygotic division. This observation accounts for the close localization between these two types of pronuclei.