Cross reaction characteristics of recombinant fusion protein of human papillomavirus 16 type L2 / 中华检验医学杂志

ABSTRACT

Objective To investigate the cross reaction characteristics of recombinant human papillomavirus 16 type L2 full-length fusion protein in HPV types of 6, 11, 18.Methods The serum samples of 108 condyloma acuminatum patients, 156 cervix cancer patients and 100 healthy control subjects were collected.The gene of full-length HPV16 L2 was amplificated from the tissue DNA of cervical cancer patient and inserted into the prokaryotic expression vector PGEX-4T-1 to construct the recombinant plasmid PGEX-4T-1-HPV16 L2.After sequencing identification, the recombinant plasmid was tranformed into E.coli BL21 (DE3).After induction by IPTG, the fusion protein containing HPV16 L2 was expressed and analyzed by both SDS-PAGE and WB.Furthermore, the specific binding capacity of the fusion protein to the HPV 6,11, 16 and 18 DNA positive patient's sera were analyzed by WB.The fusion protein was purified with NiNTA Agarose Kit and coated with ELISA reaction plates.The specific serum IgG were detected by indirect ELISA.Results The recombinant plasmid PGEX-4T-1-HPV16 L2 was constructed successfully. Highly expressed HPV16 L2 full-length fusion protein was obtained and the expression level was 27.2 %.The relative molecular mass(Mr) of the fusion protein is about 82 × 103, which matches up to the expected Mr.Meanwhile, the sera of HPV 6,11,16,18 DNA positive patients were used as the primary antibody and the Mr of the specific band was detected to be about 82 × 103 by WB.The results of indirect ELISA showed that the average levels of specific IgG in condyloma acuminatum group, cervical cancer group and healthy control subjects were 0.848 ±0.257, 0.822 ±0.247 and 0.173 ±0.143 with the positive rate of 92.6%, 94.2%and 8.0% respectively.There was no significance of the specific IgG levels between condyloma acuminatum group and cervical cancer group ( F = 0, P ＞ 0.05 ), but there was significant difference of specific IgG levels and positivity among the three groups ( F = 305.201 ,x2 = 253.178, P ＜ 0.01 ).Conclusions The HPV16 L2 full-length fusion protein has better antigenicity.However cross reactions with HPV6, 11 and 18 were found.It can be applied in serological screening reagents for HPV infection and associated cancer.