Immunotoxicological assay using fish neutrophil and toxic effects of the water pollutants

1.17-β雌二醇对罗非鱼中性粒细胞吞噬功能的影响：性别差异研究我们的评估方法不受鱼类性别的影响，这是一个很大的优势，因为不需要根据性别来选择测试鱼。2.从罗非鱼的鱼囊和头肾分离的两个细胞群的吞噬活性比较结果显示，鱼囊中性粒细胞的结果比头肾的细胞更一致和重复性更好。这是因为鱼囊中的细胞组成和成熟更为均匀。3.用原位杂交法观察雌雄鱼中性粒细胞雌激素受体(ER)α和β的表达以及ERα和ERβ的表达。中性粒细胞(1.0×10~(-6)细胞/50μL)用雌二醇(100 Ppb)处理2小时。2)加入100μL共轭亚油酸(40μM)、300μL和50μL(μg/ml)。3)混合溶液，立即测量累积信号15分钟。在28℃。这个方案可以量化从暴露的中性粒细胞到雌二醇的化学发光强度。

1.Study of tilapia neutrophil phagocytic activities after exposure to 17-beta estradiol (E2) : sensitivity in sexual differencesNeutrophil phagocytic activities were statically depressed after exposure to E2. Our appraisal method was not influenced by sex of the fish and this is a great advantage because it is not necessary to select the test fish depending on the their sex.2.Comparison of phagocytic activities between two cell populations isolated from the swim bladder and head kidney of tilapiaThe swim bladder neutrophil showed more consistent and repeatable results than the cells from the head kidney. This is because cell composition and maturation were more homogenous in the swim bladder. Isolation of cells from the swim bladder was easier than those from the head kidney.3.Expression of estrogen receptor (ER) α and βExpression of ER α and ER β were observed in neutrophil from both male and female fish using the in situ hybridization method. In the red blood cell, no signal was seen in both sexes.4.Protocol design to detect super oxide anion from the neutrophil1)Neutrophils (1.0 x 10^6 cells/50μl) were treated with the E2 (100 ppb) for 2 hrs.2)100μl of CLA(40μM), 300μl of PBS and 50μl of PMA(μg/ml) were added.3)The solution was mixed and the accumulating signal immediately was measured for 15 min. at 28℃.This protocol can quantify the intensity of chemiluminescence from exposed neutrophils to E2.