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| 结合条件对羊抗兔抗体连接到细菌磁小体的影响 | |
| 引用本文： | 陈继峰,李颖,姜伟,王珍芳,孙建波,李健,李季伦,李绍华.结合条件对羊抗兔抗体连接到细菌磁小体的影响[J].农业生物技术学报,2006,14(3):423-428. |
| 作者姓名： | 陈继峰 李颖 姜伟 王珍芳 孙建波 李健 李季伦 李绍华 |
| 作者单位： | 1. 中国农业大学微生物系,北京,100094;中国科学院植物研究所,北京,100093;郑州大学生物工程系,郑州,450001 2. 中国农业大学微生物系,北京,100094 3. 中国科学院植物研究所,北京,100093 |
| 摘 要： | 用羊抗兔抗体进行抗体连接到细菌磁小体(BMPs)实验，以探讨结合条件对抗体连接效率(以每毫克BMPs上连接抗体的微克数表示，单位为μg/mg)的影响。实验表明，冷冻干燥后的BMPs的抗体连接效率降低36%~55%；在液氮中(-196 ℃)预处理的抗体连接效率显著高于在-70 ℃和-20 ℃预处理的。室温(15 ℃)下反应18 h的抗体连接效率最高，为64.06 μg/mg，明显高于其它温度和连接时间的。反应体系中，BMPs用量由5 mg降到0.1 mg，抗体用量由10 μg增加到300 μg时，抗体连接效率逐渐上升，当BMPs用量为0.1 mg，抗体用量为300 μg时，抗体连接效率达762.37 μg/mg。PBS、HEPES、Tris-HCl、MOPS、VBS以及磷酸盐等溶液可用于抗体连接到BMPs体系中，在通常使用的缓冲液pH和浓度下，几种溶液的抗体连接效率变化范围为46.28±0.58 ~ 90.83±1.64 μg/mg；并且不同的溶液有相应的使抗体连接效率达到最高时的pH和溶液浓度，当HEPES的pH为3.5，浓度为20 mmol / L；Na3PO4的pH为5.6，浓度为5 mmol / L时，抗体连接效率分别为108.01和76.78 μg/mg。对连接到BMPs上的抗体进行SDS-PAGE定性检测结果表明，抗体连接到了BMPs的膜上。 |
| 关 键 词： | 细菌磁小体 羊抗兔抗体 结合条件 抗体连接效率 |
| 文章编号： | 1006-1304（2006）03-0423-06 |
| 收稿时间： | 2005-4-30 |
| 修稿时间： | 2005-8-9 |
Effects of Conjugating Conditions on Goat Anti-rabbit Antibody Immobilization onto Bacterial Magnetic Particles |
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| CHEN Ji-feng,LI Ying,JIANG Wei,WANG Zheng-fang,SUN Jian-bo,Li Jian,LI Ji-lun,LI Shao-hua.Effects of Conjugating Conditions on Goat Anti-rabbit Antibody Immobilization onto Bacterial Magnetic Particles[J].Journal of Agricultural Biotechnology,2006,14(3):423-428. | |
| Authors: | CHEN Ji-feng LI Ying JIANG Wei WANG Zheng-fang SUN Jian-bo Li Jian LI Ji-lun LI Shao-hua |
| Abstract: | Goat anti-rabbit immunoglobulin G (antibody) was applied as a sample for studying antibody conjugation onto bacterial magnetic particles (BMPs). The conditions for this study were different pre-treatment BMPs (-196 ℃, -70 ℃ and -20 ℃ for 5 h) for freeze-dry, temperatures with varied conjugation durations, amount of BMPs and antibody, conjugation buffers, and buffer pH and concentrations. The results showed that different pre-treatments for freeze-dry BMPs had different linkage rate of antibody (LRA). Liquid nitrogen(-196 ℃) as a pre-treatment for 5 h, the LRA was significantly higher than that of -70 ℃ and -20 ℃ pre-treatments. Any kinds of pre-treatments and freeze-dry BMPs, the LRAs were decreased 36%~55% compared with no freeze-dry BMPs. Room temperature (about 15 ℃), and incubation for 18 h resulted in the highest LRA (64.06 μg/mg) in the applied temperatures and conjugation durations. LRAs depended upon the amount of antibody and BMPs, when BMPs decreased from 5.0 mg to 0.1 mg, and antibody increased from 10 μg to 300 μg, the LRAs increased gradually. When BMPs at 0.1 mg /mL and antibody at 300 μg/mL were used, the LRA reached to 762.37 μg/mg. Many buffers could be used for antibody conjugation onto BMPs, and under the normal buffer pH and concentration, the LRAs of 8 buffers were 46.28±0.58 ~ 90.83±1.64 μg/mg. The different buffer had different pH and concentration to reach the peak LRA. When pH 3.5 and 20 mmol / L for HEPES, pH 5.6 and 5 mmol / L for Na3PO4, the LRAs were 108.01 and 76.78 μg/mg respectively. SDS-PAGE showed that antibody was conjugated onto BMPs. |
| Keywords: | bacterial magnetic particles goat anti-rabbit immunoglobulin G conjugation condition linkage rate of antibody |
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