乙肝病毒核心蛋白病毒样颗粒表面抗原密度对抗体应答水平的影响

Correlation between antigen density on the surface of hepatitis B virus core protein virus-like particles and their antibody response

【目的】为了探究乙肝病毒核心蛋白(HepatitisBviruscoreprotein,HBc)病毒样颗粒(Virus-like particles,VLPs)表面抗原密度对免疫后抗体应答水平的影响,制备了不同抗原密度的HBc VLPs疫苗,并检测了其在小鼠体内的抗体应答水平。【方法】首先制备了N端带有3个甘氨酸的人巨细胞病毒重组抗原域AD-4作为模式抗原,接着通过Sortase A的介导将AD-4连接到HBc VLPs表面上。将系列浓度梯度AD-4抗原在SortaseA介导下分别与相同浓度的HBcVLPs发生反应,制备不同抗原密度的HBc-AD-4 VLPs。将其分别免疫6–8周龄BALB/c小鼠3次,每次免疫间隔2 w,间接ELISA法检测被免疫小鼠血清的抗体应答水平。【结果】结果表明,当HBc VLPs表面抗原密度为44.4%时,即HBc反应浓度∶AD-4反应浓度为1:0.5时,不足以引起高滴度的抗体产生;当HBc VLPs表面抗原密度为64.2%时,即HBc反应浓度∶AD-4反应浓度为1:1时,HBc-AD-4 VLPs诱导的AD-4特异性抗体滴度与100%抗原密度的HBc-AD-4VLPs所引起的抗体滴度相当;当HBcVLPs表面抗原密度大于64.2%时,引起的抗体应答水平不因抗原密度增加而进一步增强。【结论】发现了HBcVLPs表面抗原密度与免疫后抗体滴度呈正相关,然而免疫64.2%抗原密度的HBc VLPs所产生的抗体滴度可达峰值,抗原密度进一步增加,抗体应答水平不会进一步加强。

[Objective]To better understand how antigen density on the surface of hepatitis B virus core protein(HBc)virus-like particles(VLPs)affects their antibody response,we generated HBc VLPs with different antigen densities,and tested their antibody response in mice.[Methods]First,we prepared the recombinant antigen domain 4(AD-4)of human cytomegalovirus(HCMV)as a model antigen,which contains three glycine molecules at its N-terminus,for Sortase A-mediated ligation onto the HBc VLPs.Displaying different densities of antigens onto the surface of VLPs was achieved by using a serial diluted recombinant AD-4 in ligation reactions.After that,HBc-AD-4 VLPs with different antigen densities were applied to 6-8 weeks old BALB/c mice.Each group was inoculated three times at 2-week intervals and the AD-4-specific IgG was detected by indirect ELISA.[Results]When the reaction concentration ratio between HBc and AD-4 is 1:0.5,which HBc surface antigen density is 44.4%,VLPs couldn’t induce a high antibody titer.When the reaction concentration ratio between HBc and AD-4 is 1:1?which HBc surface antigen density is 64.2%,VLPs could induce similar highest humoral immune response compared to 100%antigen density HBc VLPs.When HBc surface antigen density is greater than 64.2%,no further enhancement of antibody response was observed by further increasing the antigen density.[Conclusion]In conclusion,we found that antigen density on HBc VLPs is positively correlated with the antibody response.However,it reaches a peak at 64%antigen density,and no further enhancement of antibody response was observed by further increasing the antigen density.