纳米孔技术对PD-1抗原抗体结合检测的实验研究

doi:10.6023/A18110472

化学学报 ›› 2019, Vol. 77 ›› Issue (3): 287-292.DOI: 10.6023/A18110472 上一篇

研究论文

纳米孔技术对PD-1抗原抗体结合检测的实验研究

傅方舟^a,b, 张志诚^a,b, 孙倩怡^a,b, 徐冰^a,b, 沙菁㛃^a,b

a 东南大学江苏省微纳生物医疗器械设计与制造重点实验室南京 211189;
b 东南大学机械工程学院南京 211189

投稿日期:2018-11-22 发布日期:2019-01-18
通讯作者: 沙菁? E-mail:major212@seu.edu.cn
基金资助:
项目受国家自然科学基金（Nos.51675101，51435003，51375092）和江苏省研究生科研创新计划项目（SJCX18_0019）资助.

Label-free Detection of PD-1 Antibody and Antigen Immunoreaction Using Nano-Sensors

Fu Fangzhou^a,b, Zhang Zhicheng^a,b, Sun Qianyi^a,b, Xu Bing^a,b, Sha Jingjie^a,b

a Jiangsu Key Laboratory for Design and Manufacture of Micro-Nano Biomedical Instruments, Southeast University, Nanjing 211189;
b School of Mechanical Engineering, Southeast University, Nanjing 211189

Received:2018-11-22 Published:2019-01-18
Supported by:
Project supported by the National Natural Science Foundation of China (Nos. 51675101, 51435003, 51375092) and the Postgraduate Research & Practice In-novation Program of Jiangsu Province (SJCX18_0019).

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摘要/Abstract

肿瘤的免疫疗法是利用人体自身的免疫系统去治疗肿瘤的一种方法，程序性死亡受体1（PD-1）是肿瘤免疫疗法中的一种免疫检查点，利用PD-1或PD-L1的单克隆抗体，可以阻断PD-1/PD-L1信号通路，恢复T细胞的免疫杀伤功能，实现肿瘤的免疫治疗.为了研究PD-1抗体药物与人体PD-1抗原的结合有效性，以PD-1蛋白质分子为实验对象，用纳米孔传感器件开展了PD-1、PD-1抗体及其结合物的辨识.分别对纯PD-1蛋白质分子和其抗体分子进行了纳米孔过孔实验研究，发现PD-1及其抗体的相对堵塞电流幅值比分别为0.00404和0.01297，实现了抗原抗体分子的区分.并对Si₃N₄纳米孔内壁进行了PD-1抗体修饰，再将PD-1蛋白质分子通过经抗体修饰后的纳米孔，以期实现结合物的检测，实验结果显示部分抗原抗体实现了特异性结合，剩下部分呈游离状，所以纳米孔技术可实现抗原抗体结合物的区分.未来，纳米孔有望成为无标志检测药物有效性的一种新手段.

关键词: 纳米孔, 程序性死亡受体1(PD-1), 抗体, 抗原, 无标志

Immunotherapy for cancer is a method to treat cancer by using the body's own immune system. Programmed death receptor 1 (PD-1) is one of the checkpoints in the immunotherapy. The signal pathway PD-1 (programmed death receptor 1)/PD-L1 (ligand of PD-1) is closely related to the immune escape of the cancer cells. The inhibitor drugs for PD-1 checkpoint, essentially the monoclonal antibodies of PD-1 or PD-L1 which is essentially the immune checkpoints inhibitors could block the PD-1/PD-L1 pathway and reactivate T-cells to kill cancer cells, and as a result, the immunotherapy for cancer is realized. In order to study the binding process of PD-1 drugs and PD-1 antigen in vivo, in this work, solid-state nanopore as a single molecule method is used to detect the binding of PD-1 antibody and antigen. The PD-1 antibody as well as antigen is driven though the same nanopore under the same experimental condition by the external electric field. Since the antibody's block is about 0.01297 while the antigen's block is 0.00404, the PD-1 antibody is distinguished with the PD-1 antigen according to the theoretical formula. Driving the PD-1 antigen though the nanopore modified by PD-1 antibody (a series of experiments are conducted for characterization) under the same temperature and buffer concentration, the antibody-antigen complexes are detected and distinguished with PD-1 protein and its antibody through the relative current drop analysis and the current drop achieved before. The results suggest that the antibody and antigen have a specific binding (the smaller peak represents the free PD-1 antibody and antigen) and the binding process can be detected by nano-sensors. So the nanopore is able to distinguish the antibody, the antigen and the complexes without any labling. And in the future, the nanopore technology may be a rapid and label-free way for patients and doctors to evaluate the drugs' efficiency.

Key words: nanopore, programmed death receptor 1 (PD-1), antibody, antigen, label-free

引用此文

傅方舟, 张志诚, 孙倩怡, 徐冰, 沙菁㛃. 纳米孔技术对PD-1抗原抗体结合检测的实验研究[J]. 化学学报, 2019, 77(3): 287-292.

Fu Fangzhou, Zhang Zhicheng, Sun Qianyi, Xu Bing, Sha Jingjie. Label-free Detection of PD-1 Antibody and Antigen Immunoreaction Using Nano-Sensors[J]. Acta Chim. Sinica, 2019, 77(3): 287-292.

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参考文献

[1] Chen, M. J.; Novaes, P. E.; Gadia, R. Rev. Assoc. Med. Bras. 2017, 63, 729.
[2] Meyer, L. A.; Cronin, A. M.; Sun, C. C. J. Clin. Oncol. 2016, 34.
[3] Salto, S.; Shimozuma, K.; Ohashi, Y. Value Health 2007, 10, A340.
[4] Riaz, S. P.; Lüchtenborg, M.; Jack, R. H. Eur. J. Cancer 2012, 48, 54.
[5] Matsuzawa, F.; Ohki, S. Y.; Aikawa, S. I. Sci. Adv. Mater. 2005, 6, 463.
[6] Saleh, O. A.; Sohn, L. L. Proc. Natl. Acad. Sci. U. S. A. 2003, 100, 820.
[7] Schibel, A. E. P.; Ervin, E. N. Langmuir 2014, 30, 11248.
[8] Wang, S.; Haque, F.; Rychahou, P. G. ACS Nano 2013, 7, 9814.
[9] Stephen, W. H.; Eric, M.; Iftekhar, A. BMC Bioinformatics 2007, 8, S20.
[10] Wu, H. J.; Li, Y.; Fan, J. Anal. Chem. 2014, 86, 1988.
[11] Brower, V. JNCI, J. Natl. Cancer Inst. 2015, 107, djv069.
[12] Li, Q.; Quan, L.; Lyu, J. Oncotarget 2016, 7, 64967.
[13] Miller, K. D.; Siegel, R. L.; Lin, C. C. CA-Cancer J. Clin. 2016, 66, 271.
[14] Muqbil, I.; Azmi, A. S.; Mohammad, R. M. Cancers 2018, 10, 138.
[15] Barbee, M. S.; Ogunniyi, A.; Horvat, T. Z. Ann. Pharmacother. 2015, 49, 907.
[16] Cao, C.; Liao, D.-F.; Ying, Y.-L.; Long, Y.-T. Acta Chim. Sinica 2016, 74, 734. (曹婵, 廖冬芳, 应佚伦, 龙亿涛, 化学学报, 2016, 74, 734.)
[17] Sha, J. J.; Xu, B.; Chen, Y. F.; Yang, Y. J. Acta Chim. Sinica 2017, 75, 11. (沙菁?, 徐冰, 陈云飞, 杨颜菁, 化学学报, 2017, 75, 11.)
[18] Liao, D. F.; Cao, C.; Ying, Y. L. Small 2018, 14, 1704520.
[19] Kasianowicz, J. J.; Bezrukov, S. M. Nat. Biotechnol. 2016, 34, 481.
[20] Feng, Y.; Zhang, Y.; Ying, C. Genom. Proteom. Bioinf. 2015, 13, 4.
[21] Howorka, S.; Cheley, S.; Bayley, H. Nat. Biotechnol. 2001, 19, 636.
[22] Wanunu, M. Phys. Life Rev. 2012, 9, 125.
[23] Liu, N. N.; Yang, Z. K.; Ou, X. W. Mikrochim. Acta 2016, 183, 955.
[24] Venkatesan, B. M.; Bashir, R. Nat. Nanotechnol. 2011, 6, 615.
[25] Ying, Y.-L.; Zhang, X.; Liu, Y.; Xue, M.-Z.; Li, H.-L.; Long, Y.-T. Acta Chim. Sinica 2013, 71, 44. (应佚伦, 张星, 刘钰, 薛梦竹, 李洪林, 龙亿涛, 化学学报, 2013, 71, 44.)
[26] Nguyen, B. H.; Nguyen, V. H. Adv. Nat. Sci.-Nanosci. 2016, 7, 023002.
[27] Stoloff, D. H.; Wanunu, M. Curr. Opin. Biotechnol. 2013, 24, 699.
[28] Han, A.; Creus, M.; Schürmann, G. Anal. Chem. 2008, 80, 4651.
[29] Freedman, K. J.; Bastian, A. R.; Chaiken, I.; Kim, M. J. Small 2013, 9, 750.
[30] Kwak, D.; Chae, H.; Lee, M. Angew. Chem., Int. Ed. 2016, 128, 5807.
[31] Ying, Y. L.; Yu, R. J.; Hu, Y. X. Chem. Commun. 2017, 53, 8620.
[32] Wang, H. F.; Huang, F.; Gu, Z. Chin. J. Anal. Chem. 2018, 46, 50. (王慧锋, 黄飞, 顾震, 分析化学, 2018, 46, 50.)
[33] Lin, Y.; Ying, Y. L.; Gao, R. Acta Chim. Sinica 2017, 75, 675. (林瑶, 应佚伦, 高瑞, 王慧峰, 龙亿涛, 化学学报, 2017, 75, 675.)
[34] Li, Q.; Lin, Y.; Ying, Y. L.; Liu, S. C.; Long, Y. T. Sci. Sin.:Chim. 2017, 47, 1445. (李巧, 林瑶, 应佚伦, 刘少创, 龙亿涛, 中国科学:化学, 2017, 47, 1445.)
[35] Deblois, R. W.; Bean, C. P. Rev. Sci. Instrum. 1970, 41, 909.
[36] Han, A. P.; Schurmann, G.; Mondin, G.; Bitterli, R. A.; Hegelbach, N. G.; de Rooij, N. F.; Staufer, U. Appl. Phys. Lett. 2006, 88, 350.
[37] Larkin, J.; Henley, R. Y.; Muthukumar, M.; Rosenstein, J. K.; Wanunu, M. Biophys. J. 2014, 106, 696.
[38] Kowalczyk, S. W.; Grosberg, A. Y.; Rabin, Y. Nanotechnology 2011, 22, 315101.
[39] Dekker, C. Nat. Nanotechnol. 2007, 2, 209.
[40] Gierak, J.; Madouri, A.; Biance, A. L. Microelectron. Eng. 2007, 84, 779.
[41] Ma, J.; Qiu, Y.; Yuan, Z. Phys. Rev. E 2015, 92, 022719.

纳米孔技术对PD-1抗原抗体结合检测的实验研究

Label-free Detection of PD-1 Antibody and Antigen Immunoreaction Using Nano-Sensors

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