OBSERVATIONS ON THE PRIMARY CULTURE OF HEMOCYTES OF PENAEUS

R. D. Ellender, University of Southern Mississippi
AK NAJAFABADI
Bobby L. Middlebrooks, University of Southern Mississippi

Abstract

A study of the primary culture of hemocytes of Penaeus was conducted to examine growth and maintenance of these cellular populations. Hemocytes rapidly attached to flask surfaces and could be maintained for approximately one month. A medium consisting of 2 x L-15, 20% Cellect Gold fetal calf serum, 20% AKN salt solution, and other supplements was suitable for long term hemocyte maintenance. Initial cell counts were found to be more representative of monolayer confluency than volume of hemolymph collected. Bacterial contamination of cell cultures was common, but could be controlled by the addition of antibiotics to the growth medium. Contaminants were identified as members of the genera Vibrio, Pseudomonas, Flavobacterium, Aeromonas, or Proteus. Attempts to subculture hemocyte monolayers using enzymatic and mechanical methods were unsuccessful. Additional investigations concluded that hemocyte cryoprotection by DMSO was poor when cells were frozen at -20 or -70-degrees-C, that monolayer integrity was optimal at a temperature range of 25-32-degrees-C, that hemocyte populations appear to contain heterologous surface residues, and that only 1-2% of fresh hemocytes are in an active proliferation cycle at the time of collection.