本文選題：馬來絲蟲　切入點(diǎn)：肌球蛋白表位基因　出處：《中國(guó)寄生蟲學(xué)與寄生蟲病雜志》2017年01期 　論文類型：期刊論文

【摘要】：目的觀察馬來絲蟲肌球蛋白29(Brugia malayi myosin 29,Bm M29)表位基因真核表達(dá)質(zhì)粒pc DNA3.1(+)-Bm M29和原核表達(dá)的純化重組蛋白r Bm M29免疫小鼠誘導(dǎo)的免疫應(yīng)答效果。方法在大腸埃希菌(Escherichia coli)BL21誘導(dǎo)表達(dá)重組蛋白r Bm M29,純化后作為重組蛋白疫苗;純化真核表達(dá)重組質(zhì)粒pc DNA3.1(+)-Bm M29作為核酸疫苗。核酸疫苗免疫注射部位為小鼠后腿脛前肌,重組蛋白為皮下多點(diǎn)注射。60只BALB/c小鼠隨機(jī)分為A、B、C、D、E組,每組12只,分別注射PBS(100μg)、pc DNA3.1(+)/Cp G(100μg/30μg)、pc DNA3.1(+)-Bm M29/Cp G(100μg/30μg)、r Bm M29/Cp G(50μg/30μg)、pc DNA3.1(+)-Bm M29/r Bm M29/Cp G(前2次注射pc DNA3.1(+)-Bm M29/Cp G 100μg/30μg,第3次注射r Bm M29/Cp G 50μg/30μg),共免疫3次,每次免疫間隔2周。初次免疫后第4、6、8周,眼球摘除法采血制備血清,ELISA法檢測(cè)免疫小鼠血清特異性Ig G抗體效價(jià)。免疫第8周處死小鼠,制備脾細(xì)胞懸液培養(yǎng)48 h,ELISA檢測(cè)培養(yǎng)上清中細(xì)胞因子γ干擾素(IFN-γ)、白細(xì)胞介素-4(IL-4)水平。結(jié)果 ELISA檢測(cè)結(jié)果顯示,A、B、C、D和E組小鼠初次免疫后第4周血清中抗體的吸光度(A490值)分別為0.038±0.050、0.053±0.009、0.360±0.035、0.456±0.025、0.370±0.025,第6周分別為0.045±0.003、0.045±0.005、0.510±0.018、0.548±0.010、0.552±0.018,第8周分別為0.041±0.004、0.044±0.009、0.606±0.047、0.674±0.042、0.770±0.041,C、D、E組均高于A、B組(P0.05),第8周時(shí),E組Ig G抗體的A490值高于C組和D組(P0.05)。初次免疫后第8周,A至E組小鼠脾細(xì)胞培養(yǎng)上清中IFN-γ的水平分別為(47.72±8.94)、(50.43±2.81)、(304.78±8.42)、(242.28±5.99)、(426.52±6.76)pg/ml,C、D、E組均高于A、B組(P0.05),E組高于C組和D組(P0.05);小鼠脾細(xì)胞培養(yǎng)上清中IL-4的水平分別為(60.00±11.14)、(57.71±15.95)、(93.17±12.56)、(96.67±11.48)、(101.17±5.81)pg/ml,C、D、E組均高于A、B組(P0.05)。結(jié)論 pc DNA3.1(+)-Bm M29核酸疫苗和r Bm M29重組蛋白均可誘導(dǎo)BALB/c小鼠產(chǎn)生特異性體液和細(xì)胞免疫應(yīng)答,核酸疫苗與重組蛋白疫苗聯(lián)合免疫效果更佳。
[Abstract]:Objective to observe the immune response induced by eukaryotic expression plasmid pc DNA3.1 (pc BmM29) of myosin 29 Brugia malayi myosin 29 (BmM29) and recombinant protein rBmM29 of E. coli Escherichia coli. Methods the recombinant protein rBmM29 was used to immunize mice with the recombinant protein rBmM29. Methods the recombinant protein rBmM29 was used to immunize mice with Escherichia coli)BL21. The recombinant protein rBmM29 was induced to express and purified as a recombinant protein vaccine. The recombinant eukaryotic expression plasmid pc DNA3.1 was purified. The injection site of nucleic acid vaccine was the anterior tibial muscle of the hind leg of mice, and the recombinant protein was injected subcutaneously. 60 BALB/c mice were randomly divided into 2 groups, 12 in each group. Pc DNA 3.1 (Bm M29r / Bm M29Cp / 100 渭 g / 30 渭 g / 30 渭 g / 100 渭 g / 30 渭 g / 100 渭 g / 30 渭 g / 30 渭 g / 30 渭 g / 30 渭 g / 30 渭 g / 30 渭 g / 30 渭 g / 30 渭 g / 30 渭 g / 30 渭 g / g, 50 渭 g / 30 渭 g / g, respectively) (Bm / M29r / Bm M29cp / 100 渭 g / 30 渭 g / g of pc DNA) (Bm M29r / M 29Cp / 100 渭 g / g / 30 渭 g / g, third injection of r Bm M29Cp / G 50 渭 g / 30 渭 g / g / 30 渭 g / g), 2 weeks after first immunization, 468 weeks after first immunization, 4 渭 g / g / 30 渭 g / g of pc DNA (Bm M29r / Bm M29Cp / 100 渭 g / 30 渭 g / g) respectively. Serum prepared by enucleation of eyeball was used to detect the specific IgG antibody titer of immunized mice by Elisa. The mice were killed at the 8th week after immunization. The levels of IFN- 緯 and IL-4 in the supernatant were detected by Elisa. Results the results of ELISA showed that the absorbance of the antibody in serum of the mice in group E and group A was 4 weeks after the first immunization, and the results of Elisa showed that the levels of IFN- 緯 and IL-4 in the supernatant of the supernatant were determined by Elisa. Results the results of ELISA showed that the antibody in the serum of the mice in the first immunization group was A490. The values were 0.038 鹵0.050, 0.053 鹵0.009, 0.360 鹵0.035, 0.456 鹵0.025, 0.370 鹵0.025, 0.045 鹵0.0030.045 鹵0.005, 0.510 鹵0.018, 0.548 鹵0.010, 0.552 鹵0.018, 0.041 鹵0.0040.044 鹵0.009, 0.606 鹵0.047, 0.674 鹵0.0420.770 鹵0.0420.7 鹵0.041, respectively. At the 8th week, the A490 value of IgG antibody in E group was higher than that in C group and D group. 8 weeks after primary immunization, the A490 value of IgG antibody in E group was higher than that in E group. The levels of IFN- 緯 in the culture supernatant of mice were 47.72 鹵8.94, 50.43 鹵2.81, 304.78 鹵8.42, 242.28 鹵5.9921, 426.52 鹵6.76pgml / ml, 426.52 鹵6.76, respectively, and the levels of IL-4 in the culture supernatant of spleen cells in mice were higher than those in group A and B, respectively. Conclusion the levels of IL-4 in the culture supernatant of spleen cells of mice were 60.00 鹵11.147.71 鹵15.957.71 鹵15.957.17 鹵11.48A, respectively. Induce specific humoral and cellular immune responses in BALB/c mice. The combined immunization effect of nucleic acid vaccine and recombinant protein vaccine is better.
【作者單位】： 南通大學(xué)醫(yī)學(xué)院病原生物學(xué)系;
【基金】：南通市應(yīng)用研究計(jì)劃項(xiàng)目(No.MS12015115) 南通大學(xué)大學(xué)生創(chuàng)新訓(xùn)練計(jì)劃項(xiàng)目(No.yxy201509)~~
【分類號(hào)】：R392

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