抗幽门螺杆菌重组VacA卵黄抗体保护剂的探讨

Research on protectant for immunoglobulin yolk against the vacuolization cytotoxin antigen of Helicobacter pylori

目的比较硫糖铝和多种糖类对抗幽门螺杆菌重组VacA卵黄抗体保护作用,为制备能耐受胃酸和蛋白酶的IgY制剂奠定基础。方法诱导DH5-αvacA-pQE30大量表达VacA蛋白,制备纯化的VacA IgY。在不同酸性的IgY液中加入同浓度各种保护剂,在相同酸性的IgY液中加入不同浓度的各种保护剂,然后加入不同浓度硫糖铝和正常胃内浓度或高浓度的胃蛋白酶,以ELISA法测定试验组IgY的剩余抗体活性（AR）。结果在pH2.0和3.0条件下,各保护剂均能提高IgY抗体活性,且硫糖铝对IgY抗体活性的保护作用优于糖类,在此条件下50%和40%硫糖铝均可完全保持IgY抗体活性。在pH2.0-3.0及正常或高胃蛋白酶浓度下,30%以上硫糖铝均可有效保护IgY的活性。结论低pH下硫糖铝对IgY抗体活性的保护作用优于糖类,且30%以上硫糖铝可增强VacA IgY对胃蛋白酶的耐受能力,是较理想的IgY保护剂。

Objective To compare the protective effect of sucralfate on activity of immunoglobulin yolk（IgY） with that of glucose,glucosan,sucrose,alantin,and sorbitol in order to build a base for the preparation of IgY against gastric acid and pepsin. Methods Protein VacA was expressed in DH5α-vacA-pQE30,and vacA IgY was prepared and purified.Various protectants at the same concentration were added to VacA IgY under different pH levels（2.0-3.0）;protectants at different concentrations were added to VacA IgY under the same pH（2.0）.Pepsin in normal or abnormal concentrations and sucralfate in various concentrations were added;residual antibody activity of IgY（AR）was determined by ELISA.Results Under pH 2.0 and 3.0,all of the protectant increased the activity of VacA IgY and the AR of IgY containing sucralfate was higher than all of the other carbohydrate protectants.Under pH2.0 and 3.0,50% and 40% sucralfate maintained its activity.Under pH2.0-3.0 and normal or abnormal pepsin concentrations,sucralfate above 30% effectively protected its activity.Conclusion Under a low pH,the protective effect of sucralfate on VacA IgY was better than that of the carbohydrate group.Furthermore,30% or more sucralfate improved the tolerance of VacA IgY against low pepsin,which indicates that sucralfate may be a satisfactory protectant for IgY.