PHENOLOXIDASE CHARACTERIZATION IN VACUOLAR HEMOCYTES FROM THE SOLITARY ASCIDIAN STYELA-PLICATA

Phenoloxidase (PO) activity was shown in lysates of Styela plicata hemocytes assayed spectrophotometrically by means of L-Dopa oxidation without divalent cations. Trypsin and chymotrypsin pretreatment and preincubation with microbial lipopolysaccharides significantly activated PO, whereas laminarin or zymosan were ineffective. Soybean trypsin inhibitor, tropolone, and phenylthiourea, but not benzamidine, were inhibitors. Finally, hemocytes were separated by a discontinuous Percoll density gradient to determine which cells were active. PO activity was demonstrated, by both biochemical and cytochemical assays, in the separated fraction enriched mainly with the globular granulocytes called morula cells. (C) 1995 Academic Press, Inc.