Production of polyclonal antibodies against the tegument of sparganum (plerocercoid of Spirometra mansoni) and its immunolocalization

Abstract

In a previous study, the author developed a method for separation of the tegument of spargana (plerocercoids of Spirometra mansoni) from the parenchyme using urea. The present study, as a next step, was performed to evaluate which molecules are present in the outer tegument. Two major proteins, 180 and 200 kDa, are present in the tegument and we could make polyclonal antibodies against these molecules. Their immunolocalization was processed and the outermost layer of the spargana showed strong positive staining. Conclusively, we could confirm that the 180 and 200 kDa molecules might be tightly bound membrane proteins in the tegument of spargana.

Keywords: immunohistochemistry; polyclonal antibody; sparganum; syncytial layer.

Fig. 1

7.5-15% SDS-PAGE of tegumental proteins of spargana and production of polyclonal antibodies. (A) Note the 60 kDa major single band in PBS extracts of the tegument (lane 1) and numerous molecules by detergent extracted proteins (lane 2) compared with soluble crude extracts (lane 3) of the sparganum. (B) Polyclonal antibodies against detergent extracted proteins were reacted with 2 major bands of 180 and 200 kDa molecules.

Fig. 2

Immunohistochemistry by polyclonal antibodies against tegumental molecules in spargana. Strong positive reactions are observed in the outer tegument of a sparganum (arrow). No other organelles, including calcareous corpuscles (*), show a positive reaction. (A) negative control. × 100 magnification; (B) and (C): stain with polyclonal antibodies. × 100 and × 200 magnifications, respectively (Scale bar=100 µm).