improved method of cell culture system from eye stalk, hepatopancreas, muscle, ovary, and hemocytes of Penaeus vannamei

• Published in: In vitro cellular & developmental biology. Animal 2010-10, Vol.46 (9), p.801-810
• Main Authors: George, Sunil K, Dhar, Arun K
• Format: Article
• Language: English
• Subjects: Animal Genetics and Genomics; Animals; Biomedical and Life Sciences; Carbon Dioxide - chemistry; Cell Biology; Cell Culture; Cell culture techniques; Cell Culture Techniques - methods; Cell growth; Cultured cells; Developmental Biology; Female; Hemocytes; Hemocytes - cytology; Hemolymph; Hepatopancreas; Hepatopancreas - cytology; LETTER TO THE EDITOR; Life Sciences; Lipopolysaccharides; Litopenaeus vannamei; Muscles; Muscles - cytology; Ovaries; Ovary - cytology; Penaeidae - cytology; Posterior Eye Segment - cytology; Primary cell culture; Shrimp; Specific Pathogen-Free Organisms; Stem Cells; Temperature
• ISSN: 1071-2690; 1543-706X
• DOI: 10.1007/s11626-010-9343-x

Improved methods of cell culture from eye stalk, hepatopancreas, muscle, ovary, and hemocytes of shrimp (Penaeus vannamei) were established using synthetic media and shrimp muscle extract (SME). For hemocytes and ovarian cell cultures, Grace's insect medium supplemented with 10% (v/v) fetal bovine serum and 10% SME (v/v) showed enhanced attachment and proliferation of the cells. The hemocyte and ovarian cell cultures could be maintained for 48 and 66 days, respectively, and have been sub-cultured four and six times, respectively. Both ovary and hemocyte cell cultures contained primarily epithelial-like cells. Cells derived from ovary tissue grew preferably between 26°C and 28°C with 5% CO₂. Although the temperature preference of hemocyte cells was the same as ovarian cells, CO₂ supplementation did not show any difference in the growth of hemocyte cells. When the shrimp were injected with lipopolysaccharide (8 μg/g of shrimp) and hemolymph was drawn 24 h post-injection, the in vitro multiplicity of hemocytes dramatically improved. The growth of eye stalk, hepatopancreas, and muscle-derived cells was much less compared to ovarian cells and hemocytes under the conditions described above. The optimal culture conditions for ovarian cells and hemocytes were also different from that for eye stalk, hepatopancreas, and muscle cell culture. The proliferation efficiencies of primary cultures of hepatopancreas, eyestalk, and muscle cells were about 30, 12, and