Commercially available culture media and supplements were tested for their potential to produce primary cell cultures from tissues of Indian mud crabs Scylla serrata. Eight commercially available culture media from Sigma-Aldrich (Leibovitz's L-15, Medium 199, Grace's Insect Medium, Minimal Essential Medium, Dulbecco's Modified Eagle Medium, TC-100 Insect Medium, IPL-41 Insect Medium, and Roswell Park Memorial Institute) were examined. Three different supplements (amino acid and sugar [AS], crab muscle extract [CME], and natural seawater [NSW]) were also examined. The hemocyte culture appeared to grow well for a maximum period of 21 d in 2 × L-15 medium supplemented with AS and 15% fetal bovine serum (FBS). Partial amplification and sequencing of the cytochrome oxidase subunit I (COI) gene confirmed that the primary hemocytes originated from Indian mud crabs. The effects of four metals on hemocyte viability were evaluated using the MTT assay. Of the four metals examined (arsenic, lead, cobalt, and nickel), cobalt and nickel were more toxic to the crab cells than the other metals. Both acridine orange/ethidium bromide and Hoechst staining showed the presence of apoptosis and necrosis in metal-treated groups, which suggests that metals in an aquatic environment induce death of the Indian mud crab's hemocytes. The hemocyte primary cell culture was also used to study the cytotoxicity effect of bacterial extracellular products from Vibrio harveyi and white spot syndrome virus. This study demonstrates that hemocyte primary cell culture can be used as a tool to study viral and bacterial pathogenesis and to assess the cytotoxicity of pollutants present in aquatic environments.

中文翻译：

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印度泥蟹 Scylla serrata 细胞培养系统的开发和表征

测试了市售培养基和补充剂从印度泥蟹 Scylla serrata 组织产生原代细胞培养物的潜力。来自 Sigma-Aldrich 的八种市售培养基（Leibovitz's L-15、Medium 199、Grace's Insect Medium、Minimal Essential Medium、Dulbecco's Modified Eagle Medium、TC-100 Insect Medium、IPL-41 Insect Medium 和 Roswell Park Memorial Institute）审查。还检查了三种不同的补充剂（氨基酸和糖 [AS]、蟹肉提取物 [CME] 和天然海水 [NSW]）。血细胞培养物似乎在补充有 AS 和 15% 胎牛血清 (FBS) 的 2 × L-15 培养基中生长良好，最长可达 21 天。细胞色素氧化酶亚基 I (COI) 基因的部分扩增和测序证实原代血细胞来源于印度泥蟹。四种金属对血细胞活力的影响使用 MTT 测定法进行评估。在检查的四种金属（砷、铅、钴和镍）中，钴和镍对蟹细胞的毒性比其他金属更大。吖啶橙/溴化乙锭和 Hoechst 染色均显示金属处理组中存在细胞凋亡和坏死，这表明水环境中的金属会诱导印度泥蟹血细胞的死亡。血细胞原代细胞培养物还用于研究来自哈维弧菌和白斑综合征病毒的细菌细胞外产物的细胞毒性作用。