生物靶标光敏剂对鸭乙肝病毒的特异结合及杀灭作用研究

The specific combination and inactivation function of bio-photosensitizer target on duck hepatitis virus B

目的 :利用生物靶标效应的原理 ,探索一种特异灭活病毒的光化学方法。方法 :将针对鸭乙肝病毒 (DHBV)设计并人工合成的两条三螺旋结构寡核苷酸 (TFO)分别标记 8-甲氧基补骨脂素 (8-MOP) ,并将这种复合物 (TFO -P)与长波紫外线 (UVA)共同作用于血液中的DHBV ,通过原代细胞感染、酶联免疫测定、DNA斑点杂交等试验 ,观察在一定作用条件下 ,TFO -P对DHBV-DNA的特异结合及其所产生的特异光敏效应对DHBV的灭活程度。结果 :DHBV -DNA样品斑点印迹随TFO -P含量的增加明显增强 ,而同时处理的血液中的白细胞DNA样品斑点印迹无明显增强 ;在TFO -P的加入量为 0 1nmol/ml,UVA照射强度为180 0 μW /cm2 时 ,5～ 2 0min的UV照射可灭活DHBV 1 90～ 6 4个对数级 ,灭活病毒的效率显著高于UVA和 8-MOP的单独作用时的效果。结论 :TFO -P与血液中DHBV -DNA能特异结合 ,在适宜的处理条件下 ,所产生的光敏效应能有效灭活血液中的DHBV ,灭活滴度可达

Objective:To explore a photochemical method that can inactive virus specifically by based on the principle of bio-photosensitizer which having the target effects.Method:The triple helix forming oligonucleotide(TFO) was labelled with 8-methoxypsoralen(8-MOP) at 3′ end.When in coordination with long wave ultraviolet ray(UVA),this decorated complex(TFO-P) was added into the blood,to affect the contaminated virus--duck hepatitis virus B (DHBV).At the designed condition,the efficacy of special binding to DHBV-DNA and virual inactivation by TFO-P was detected by assays of the DNA dot blot hybridization,infection of primary culture of duck hepatocyte,and ELISA.Result:To the DHBV-DNA samples,the intensity of dot blot increased with the contents of TFO-P obviously.Otherwise,the dot blot of WBC-DNA′s did not changed with the content of TFO-P;When the added doses of TFO-P was about 0.1 nmol/ml,and irradiated by UVA at the intensity of 1800μW/cm 2 for 5 min～20min,the DHBV could be reduced about 1.90～6.40 log.The efficacy of viral inactivation by TFO-P is significantly higher than 8-MOP and TFO respectively.Conclusion:TFO-P could specially combine to DHBV-DNA in blood.At a suitable condition,the photochemical effects of TFO-P could reduce the DHBV above the dose of 6.0 log.