Key Laboratory of Mariculture, Agriculture Ministry, PRC, Dalian Ocean University, Dalian 116023, China.
Fish Shellfish Immunol. 2010 Nov;29(5):832-8. doi: 10.1016/j.fsi.2010.07.024. Epub 2010 Jul 24.
One monoclonal antibody (mAb 3F6) against coelomocytes of sea cucumber Apostichopus japonicus was developed by immunization of Balb/C mice. Analyzed by indirect immunofluorescence assay test (IIFAT), immunocytochemical assay (ICA), Western blotting and fluorescenceactivated cell sorter (FACS), mAb 3F6 showed specific for spherulocytes of A. japonicus. The mAb 3F6 recognized an antigen of molecular weight 136 kDa in Western blotting. Isotype analysis revealed mAb 3F6 as IgG type. The flow cytometry assay confirmed the microscopy observations and showed coelomocytes positive to mAb 3F6. The antigencity of haemocytes or coelomocytes of Hemicentrotus pulcherrimus, Scapharca subcrenata, Asterina pectinifera, Asterias rollestoni, Ruditapes philipinarum, Patinopecten yessoensis and Mytilus edulis was compared and the result showed that none of them was positive with mAb 3F6. Most of cells free in polian vesicle were positive with mAb 3F6. The positive cells are in spherical shape, 5-7 microm in diameter, smaller than coelomic spherulocytes of A. japonicus. The result of immunofluorescent staining with cells in hemal vessel showed that there were strong positive signals on cytoplasm of some spherical cells with diameter of 7-8 microm. Some other cells with higher nucelo-plasmic ratio, about 5-6 microm in diameter showed weak positive signals on membrane. Immunohistochemistry assay revealed that positive signals were mostly observed in the lumen structure of rete mirabile and haemocoel of the respiratory tree. In addition, the outer epidermis of body wall and tentacle also showed positive.
一种针对海参体腔细胞的单克隆抗体（mAb 3F6）通过免疫 Balb/C 小鼠而开发。通过间接免疫荧光检测（IIFAT）、免疫细胞化学检测（ICA）、Western 印迹和荧光激活细胞分选（FACS）分析，mAb 3F6 显示对海参的球形细胞具有特异性。Western 印迹显示 mAb 3F6 识别分子量为 136 kDa 的抗原。同型分析显示 mAb 3F6 为 IgG 型。流式细胞术检测证实了显微镜观察结果，并显示体腔细胞对 mAb 3F6 呈阳性。对糙海参、中国蛤蜊、多棘海盘车、罗氏轮海星、菲律宾蛤仔、虾夷扇贝和贻贝的血细胞或体腔细胞的抗原性进行了比较，结果表明它们均与 mAb 3F6 无反应。大多数游离于体腔液中的细胞与 mAb 3F6 呈阳性反应。阳性细胞呈球形，直径为 5-7 微米，小于海参的体腔球形细胞。血管细胞免疫荧光染色结果显示，直径为 7-8 微米的一些球形细胞的细胞质上有强烈的阳性信号。其他一些核质比更高的细胞，直径约为 5-6 微米，在膜上显示弱阳性信号。免疫组织化学检测显示，阳性信号主要观察到在 rete mirabile 的腔结构和呼吸树的血腔中。此外，体壁的外表皮和触手也显示出阳性。
