Development of Multi-biomarker Assays Using Mussels for Coastal Monitoring

对单细胞电泳法（SCG）、混合功能加氧酶法和吞噬法进行了改进，并进行了室内实验和现场研究。对于Zn_2Cl暴露贻贝血细胞SCG检测的改进，最佳条件为解卷时间为15分钟，电泳时间为20分钟，其量效关系最明显。在该条件下的实验室实验中，我们可以观察到DNA损伤的剂量依赖性和时间依赖性，以及DNA损伤和修复的时间模式，表明了该方法的实用性。HPLC法测定EROD活性的灵敏度为常规方法的3000倍。采用高效液相色谱法对贻贝消化腺制备的微粒体进行分析，结果清晰、灵敏。MMS和B [a] P显著诱导了EROD活性，但未诱导ECOD活性。考察了吞噬试验的实验条件，如锥虫酚浓度、荧光大肠杆菌浓度等。随着B [a] P浓度的增加。吞噬作用被激活，在较高的浓度下被抑制。2000年8月、10月和12月在富山湾的6个港口使用这些已确定的生物标志物进行了初步的实地研究。讨论了生物标志物的测定结果。

The improvement of single cell get electrophoresis assay (SCG assay), mixed function oxygenase assay, and phagocytosis assay, laboratory experiments and field studies had been conducted. As for the improvement of SCG assay using mussel hemocytes exposed to Zn_2Cl, the optimum condition was the combination of unwinding time of 15 minutes and electrophoresis time of 20 minutes, showing the clearest dose-response relationship. In laboratory experiments with this condition, we could observe the dose-dependent and temporal DNA damage, and temporal patter of DNA damage and repair, indicating the usefulness of this assay. HPLC method for EROD activity showed the 3,000 times of sensitivity compared to the conventional method. Using HPLC method with microsome prepared from mussel digestive gland, we could get the clear and sensitive results. Significant induction of EROD activity was observed in the experiment with MMS and B [a] P, but ECOD activity had not been induced. The experimental conditions of phagocytosis assay, such as concentration of trypanbule, concentration of fluorescent E.coli etc., had been examined. With the increase of B [a] P concentration. phagocytosis had been activated, and suppressed at the higher concentration. At 6 harbors in Toyama Bay, the preliminary field study had been conducted using these established biomarkers in August, October and December 2000. The results of biomarkers were discussed.