Aquatic Animal Health Laboratory, C. Abdul Hakeem College (Autonomous), Affiliated to Thiruvalluvar University, Melvisharam, India.
J Fish Dis. 2024 May;47(5):e13927. doi: 10.1111/jfd.13927. Epub 2024 Jan 29.
A permanent cell line, SPB (Snubnose pompano brain) was established from Trachinotus blochii by the explant culture method. It has been sub-cultured more than 75 passages and showed optimal growth at 28°C using L-15 medium supplemented with 15% to 20% FBS. The SPB cells were cryopreserved at different passage levels for various applications. SPB cells were composed of fibroblastic and epithelial-like cells. The SPB cells were tested for mycoplasma contamination which was found to be negative. The origin of the SPB cell line from T. blochii was confirmed by amplification of the mitochondrial cytochrome oxidase I (COI) gene. The transfection efficiency of SPB cell line is 15% assessed by expression of green fluorescent protein using pEGFP-N1 plasmid. In addition, two CMV promotor plasmids pFNCPE42-DNA and pcDNAVP28 were transfected to SPB cells and it shows high expression levels of FNCP of fish nodavirus and VP28 protein of white spot syndrome virus by immunostaining. The SPB cells showed susceptibility to SJNNV and the infection was confirmed by RT-PCR, Western blot, ELISA, TCID and RT-qPCR. Experimental infection was carried out in T. blochii using SJNNV propagated in SPB cell line and found 100% mortality with clinical signs. The infection was confirmed by RT-PCR. The SPB cell line can be used for propagation of fish viral pathogens and production of the recombinant proteins.
从鲳鲹（Trachinotus blochii）中通过组织块培养法建立了一个永久细胞系 SPB（Snubnose pompano brain）。它已经传代培养了超过 75 代，在使用 L-15 培养基补充 15%至 20% FBS 的条件下，在 28°C 下最佳生长。SPB 细胞在不同的传代水平下被冷冻保存，用于各种应用。SPB 细胞由成纤维细胞和上皮样细胞组成。对 SPB 细胞进行支原体污染检测，结果为阴性。通过扩增线粒体细胞色素氧化酶 I（COI）基因证实了 SPB 细胞系源自鲳鲹。使用 pEGFP-N1 质粒表达绿色荧光蛋白评估 SPB 细胞系的转染效率为 15%。此外，将两个 CMV 启动子质粒 pFNCPE42-DNA 和 pcDNAVP28 转染到 SPB 细胞中，通过免疫染色显示出鱼神经坏死病毒的 FNCP 和白斑综合征病毒的 VP28 蛋白的高表达水平。SPB 细胞对 SJNNV 敏感，通过 RT-PCR、Western blot、ELISA、TCID 和 RT-qPCR 证实了感染。使用在 SPB 细胞系中繁殖的 SJNNV 在鲳鲹中进行实验感染，发现具有临床症状的 100%死亡率。通过 RT-PCR 证实了感染。SPB 细胞系可用于繁殖鱼类病毒病原体和生产重组蛋白。
