Reexamination of phenoloxidase in larval circulating hemocytes of the silkworm, Bombyx mori.
We have developed a modified method to detect phenoloxidase activity on hemocytes by using freshly prepared l-DOPA (1 mg/ml in 35% ethanol) to fix and incubate larval hemocytes. This method is more sensitive than the common method, in which hemocytes were fixed in 4% formaldehyde and then incubated with 2 mg/ml l-DOPA in water separately. Phenoloxidase assayed using this modified method can be inhibited by phenyltiourea (phenoloxidase inhibitor). After incubation with l-DOPA solution in ethanol, most prohemocytes, all plasmatocytes and young granulocytes are stained brown due to oxidation of l-DOPA into pigments, indicating that they have phenoloxidase. Oenocytoids are dimly stained because many of their cell inclusions have been released during the treatment. Large propidium-iodide-negative prohemocytes have strong phenoloxidase activity and are easily misunderstood as propidium-iodide-positive oenocytoids if the fluorescent method is not used for identification. Thus, in addition to oenocytoids and plasmatocytes, some prohemocytes and granulocytes in the silkworm also have phenoloxidase.
Ling
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Laboratory of Silkworm Physiology, Department of Applied Biology, Faculty of Textile Science and Technology, Shinshu University, Ueda, Nagano 386-8567, Japan. l_erjun@hotmail.com
Shirai
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Kanehatsu
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Kiguchi
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D000818:Animals; D012831:Bombyx / Q000166:cytology*; D006098:Granulocytes / Q000166:cytology / Q000378:metabolism; D006434:Hemocytes / Q000166:cytology / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D006651:Histocytochemistry; D007814:Larva / Q000166:cytology; D007980:Levodopa / Q000494:pharmacology; D014442:Monophenol Monooxygenase / Q000032:analysis / Q000378:metabolism*; D012680:Sensitivity and Specificity; D013194:Staining and Labeling / Q000379:methods
D007980:Levodopa; D014442:Monophenol Monooxygenase