Winotaphan Phochit, Sithigorngul Paisarn, Muenpol Oraporn, Longyant Siwaporn, Rukpratanporn Sombat, Chaivisuthangkura Parin, Sithigorngul Weerawan, Petsom Amorn, Menasveta Piamsak
Biotechnology Program, Faculty of Science, Chulalongkorn University, Bangkok 10330, Thailand.
Fish Shellfish Immunol. 2005 Mar;18(3):189-98. doi: 10.1016/j.fsi.2004.05.007.
Monoclonal antibodies specific to haemocytes of Penaeus monodon were generated from a mouse immunized with a mixture of SDS-treated and formalin-fixed haemocytes. Hybridoma clones were selected by immunohistochemistry against fixed haemocytes, heart, lymphoid organ, and haemopoietic tissue, and Western blot against haemocyte extract and haemolymph. Sixteen monoclonal antibodies specific to haemocytes were obtained and could be divided into six groups according to their binding capacities to various haemocyte proteins in Western blot analyses, 102, 43, approximately 20, 61, 175 and approximately 230 kDa, and their differences in recognition of haemocyte sub-populations. The first group of antibodies strongly recognized a small subset of semi-granulocytes (SG) and hyalinocytes (H) but occasionally stained lightly a very small population of granulocytes (G). The antibodies also bound to a group of cells in haemopoietic tissue as well as cells located at the inner layers of the tubules in the lymphoid organ but not in the spheroid. The second group of antibodies strongly bound to a large sub-population of G and SG with coarse granules but did not bind to most of the H. This group of antibodies also cross-reacted with cells in the outer layer of the tubules in the lymphoid organ. The third group of antibodies recognized all G and only a small portion of SG. The fourth, fifth and sixth groups bound to sub-populations of G, SG and H in similar proportions. None of the antibodies showed any cross-reactivity to other components in haemolymph. The common antigens recognized by the first and the second groups of antibodies in the haemopoietic tissue and the lymphoid organ may reflect relationships among these organs in the development of the sub-populations of G and SG. Haemopoietic tissue may be the site for haemocyte production and the lymphoid organ may be the site for further differentiation of at least two different lines of haemocytes.
用经十二烷基硫酸钠（SDS）处理和福尔马林固定的血细胞混合物免疫小鼠，产生了对斑节对虾血细胞具有特异性的单克隆抗体。通过针对固定血细胞、心脏、淋巴器官和造血组织的免疫组织化学，以及针对血细胞提取物和血淋巴的蛋白质印迹法筛选杂交瘤克隆。获得了16种对血细胞具有特异性的单克隆抗体，根据它们在蛋白质印迹分析中与各种血细胞蛋白（102、43、约20、61、175和约230 kDa）的结合能力以及对血细胞亚群识别的差异，可分为六组。第一组抗体强烈识别一小部分半颗粒细胞（SG）和透明细胞（H），但偶尔也会轻微染色一小部分颗粒细胞（G）。这些抗体还与造血组织中的一组细胞以及淋巴器官小管内层的细胞结合，但不与球状体中的细胞结合。第二组抗体强烈结合大部分具有粗大颗粒的G和SG亚群，但不与大多数H结合。这组抗体还与淋巴器官小管外层的细胞发生交叉反应。第三组抗体识别所有的G和仅一小部分SG。第四、第五和第六组以相似比例结合G、SG和H的亚群。没有一种抗体对血淋巴中的其他成分表现出任何交叉反应。第一组和第二组抗体在造血组织和淋巴器官中识别的共同抗原可能反映了在G和SG亚群发育过程中这些器官之间的关系。造血组织可能是血细胞产生的部位，而淋巴器官可能是至少两种不同血细胞系进一步分化的部位。
