抗原钥孔戚血蓝蛋白(KLH)对金鱼(Carassius auratus)体液免疫的影响研究

以钥孔戚血蓝蛋白(Keyhole limpet hemocyanin,KLH)为抗原、Montanide ISA 763 A为佐剂开展金鱼免疫试验,检测血清抗体水平,评估金鱼体液免疫应答情况,以期为其免疫策略的制定提供依据。试验在水温15~18℃时进行,采用(20±3)g体重的健康鱼150尾,随机均分为三组,第一组使用(KLH+Montanide ISA 763 A)免疫,第二组仅使用KLH免疫,第三组注射等量的0.01 mol/L磷酸盐缓冲液(Phosphate buffered solution,PBS)为对照,历经三次免疫并对4次采样的血清样本进行ELISA检测。结果显示,第一组和第二组均检测到较高的抗KLH抗体水平,在第二次、第三次免疫后,添加ISA佐剂的第一组抗体仍维持较高水平,而在仅用KLH免疫的第二组抗体水平下降,并降低到对照组水平,表明使用(KLH+Montanide ISA 763 A)免疫金鱼可以获得更加持久和稳定的免疫效果,而仅注射KLH的免疫效果较为短暂。研究还说明金鱼在低温情况下,可以产生良好的体液免疫应答。

Effect of keyhole limpet hemocyanin(KLH)as antigen on humoral immunity in goldfish(Carassius auratus)

In order to provide enough reference material for enacting immune strategy of goldfish(Carassius auratus),KLH was used as antigen to immunize it with adjuvant Montanide ISA 763 A,and the serum antibody level was detected to evaluate the humoral immunoresponse.The experiment was carried out under 15~18℃ and 150 healthy fish with body weight of(20±3)g were randomly divided into 3 groups.Group Ⅰ was immunized with(KLH+Montanide ISA 763A),Group Ⅱ was merely immunized with KLH and Group Ⅲ was intraperitoneal injected with equivalent PBS as control.Antiserum was sampled after using KLH plus ISA adjuvant or not from the immunized fish three times,and the antibody variation was evaluated by ELISA.As a result,the antibody level in Group Ⅰ and Group Ⅱ were high.The antibody level of Group Ⅰ was still high after the first and second booster immunization,while that of Group Ⅱ were decreased and approached to the level of Group Ⅲ.We found the fish immunized with KLH plus ISA could acquire a long and stable immunity,while the fish merely immunized with KLH could get a short and transitory one.This study also showed the goldfish possessed well humoral immunoresponse in hypothermia situation.