乳腺癌生物标志物钙网蛋白的核酸适配体筛选及血清检测和乳腺癌细胞识别

引用本文: 孙淼, 杨歌, 赵毅, 屈锋. 乳腺癌生物标志物钙网蛋白的核酸适配体筛选及血清检测和乳腺癌细胞识别[J]. 分析化学, 2020, 48(5): 642-649. doi: 10.19756/j.issn.0253-3820.201004 shu

Citation: SUN Miao, YANG Ge, ZHAO Yi, QU Feng. Screening of Aptamer for Breast Cancer Biomarker Calreticulin and Its Application to Detection of Serum and Recognition of Breast Cancer Cell[J]. Chinese Journal of Analytical Chemistry, 2020, 48(5): 642-649. doi: 10.19756/j.issn.0253-3820.201004 shu

乳腺癌生物标志物钙网蛋白的核酸适配体筛选及血清检测和乳腺癌细胞识别

北京理工大学生命学院, "分子医学与生物诊疗"工业和信息化部重点实验室, 北京 100081
基金项目:
本文系国家自然基科学金项目（Nos.21675012，21874010，21827810）资助

摘要: 钙网蛋白（CRT）与许多癌症的发生密切相关，被认为是乳腺癌侵袭和转移的有效新型生物标志物以及乳腺癌分期和预后的评价指标。本研究利用毛细管电泳-指数富集配体进化技术（CE-SELEX）筛选并鉴定了CRT的核酸适配体。毛细管电泳法、等温滴定量热法及纳米金比色法表征结果表明，适配体Apt 23对CRT具有较高的亲和力和特异性。FAM标记的Apt 23（FAM-Apt 23）可用作毛细管电泳的亲和探针，采用激光诱导荧光检测器，检测血清中CRT的浓度范围为1~1000 nmol/L，检出限为0.5 nmol/L。FAM-Apt 23也可用作CRT的免疫荧光成像探针，可结合在CRT过表达的乳腺癌细胞4T1的表面。初步研究结果表明，利用毛细管电泳筛选的核酸适配体Apt 23可用于CRT识别和检测的探针。

关键词:

English

Screening of Aptamer for Breast Cancer Biomarker Calreticulin and Its Application to Detection of Serum and Recognition of Breast Cancer Cell

School of Life Science, Beijing Institute of Technology, Key Laboratory of Molecular Medicine and Biological Diagnosis, Ministry of Industry and Information Technology, Beijing 100081, China
Received Date: 03 January 2020
Revised Date: 05 March 2020
Fund Project:
This work was supported by the National Natural Science Foundation of China (Nos. 21675012, 21874010, 21827810).

Abstract: Calreticulin (CRT) is closely related to cancer, and is considered to be a novel biomarker for invasion and metastasis of breast cancer, as well as an indicator of staging and prognosis. Here, we screened and identified aptamers of CRT for the first time with capillary electrophoresis-based systematic evolution of ligands by exponential enrichment (CE-SELEX). The high affinity and specificity Apt 23 was screened and verified by CE isothermal titration calorimetry and gold nanoparticles-based colorimetric assay. By using FAM-labeled Apt 23 (FAM-Apt 23) as an affinity probe, CRT was dected in the linear concentration range of 1-1000 nmol/L in serum by laser-induced fluorescence detector with a detection limit of 0.5 nmol/L. FAM-Apt 23 could also be used as an immunofluorescence imaging probe, which could be bound to the surface of breast cancer cells 4T1 with overexpressed CRT. Preliminary studies have shown that Apt 23 could be used as aptamer probes for recognition and detection of CRT.

Key words:

Calreticulin
/ Aptamer
/ Capillary electrophoresis based systematic evolution of ligands by exponential enrichment

1. [1]
  Smith M J, Koch G L. EMBO J.,1989,8(12): 3581-3586
2. [2]
  Païdassi H, Tacnet-Delorme P, Verneret M, Gaboriaud C, Houen G, Duus K, Ling W L, Arlaud G J, Frachet P. J. Mol. Biol.,2011,408(2): 277-290
3. [3]
  Eggleton P, Bremer E, Dudek E, Michalak M. Expert Opin. Ther. Tar.,2016,20(9): 1137-1147
4. [4]
  Chiang W F, Hwang T Z, Hour T C, Wang L H, Chiu C C, Chen H R, Wu Y J, Wang C C, Wang L F, Chien C Y, Chen J H, Hsu C T, Chen J Y. Oral Oncol.,2013,49(6): 534-541
5. [5]
  Song M N, Moon P G, Lee J E, Na M, Kang W, Chae Y S, Park J Y, Park H, Baek M C. Arch. Pharm. Res.,2012,35(10): 1839-1847
6. [6]
  Lee H H, Lim C A, Cheong Y T, Singh M, Gam L H. Int. J. Biol. Sci.,2012,8(3): 353-362
7. [7]
  Zamanian M, Hamadneh L A Q, Veerakumarasivam A, Rahman S A, Shohaimi S, Rosli R. Cancer Cell Int.,2016,16(1): 56
8. [8]
  Lwin Z M, Guo C, Salim A, Yip G W, Chew F T, Nan J, Thike A A, Tan P H, Bay B H. Modern Pathol.,2010,23(12): 1559-1566
9. [9]
  Chen C N, Chang C C, Su T E, Hsu W M, Jeng Y M, Ho M C, Hsieh F J, Lee P H, Kuo M L, Lee H, Chang K J. Ann. Surg. Oncol.,2009,16(2): 524-533
10. [10]
  Alfonso P, Núñez A, Madoz-Gurpide J, Lombardia L, Sánchez L, Casal J I. Proteomics,2005,5(10): 2602-2611
11. [11]
  Minami S, Nagashio R, Ueda J, Matsumoto K, Goshima N, Hattori M, Hachimura K, Iwamura M, Sato Y. Biomed. Res.,2014,35(1): 25-35
12. [12]
  Sheng W, Chen C, Dong M, Zhou J, Liu Q, Dong Q, Li F. J. Cell. Physiol.,2014,229(7): 887-897
13. [13]
  Alaiya A, Roblick U, Egevad L, Carlsson A, Franzén B, Volz D, Huwendiek S, Linder S, Auer G. Anal. Cell. Pathol.,2000,21(1): 1-9
14. [14]
  Hellman K, Alaiya A A, Schedvins K, Steinberg W, Hellström A C, Auer G. Brit. J. Cancer,2004,91(2): 319-326
15. [15]
  Lu Y C, Weng W C, Lee H. Biomed. Res. Int.,2015:526524
16. [16]
  Ni M, Wei W, Wang Y, Zhang N, Ding H, Shen C, Zheng F. J. Clin. Immunol.,2013,33(5): 947-953
17. [17]
  Xu Q, Chen C, Chen G, Chen W, Zhou D, Xie Y. Oncol. Lett.,2018,15(6): 8999-9008
18. [18]
  Fucikova J, Becht E, Iribarren K, Goc J, Remark R, Damotte D, Alifano M, Devi P, Biton J, Germain C, Lupo A, Fridman W H, Dieu-Nosjean M C, Kroemer G, Fridman C S, Cremer I. Cancer Res.,2016,76(7): 1746-1756
19. [19]
  Ellington A D, Szostak J W. Nature,1990,346(6287): 818-822
20. [20]
  Tuerk C, Gold L. Science,1990,249(4968): 505-510
21. [21]
  Zhou G, Wilson G, Hebbard L, Duan W, Liddle C, George J, Qiao L. Oncotarget,2016, 7: 13446-13463
22. [22]
  Mercier M C, Dontenwill M, Choulier L. Cancers,2017,9(12): 69
23. [23]
  Bayat P, Nosrati R, Alibolandi M, Rafatpanah H, Abnous K, Khedri M, Ramezani M. Biochimie,2018,154: 132-155
24. [24]
  Dong L, Tan Q, Ye W, Liu D L, Chen H F, Hu H W, Wen D, Liu Y, Cao Y, Kang J W, Fan J, Guo W, Wu W Z. Sci. Rep.,2015,5: 15552
25. [25]
  Zhu C, Li L S, Yang G, Irfan M, Wang Z J, Fang S B, Qu F. Talanta,2019,205: 120088
26. [26]
  Berezovski M, Krylov S N. J. Am. Chem. Soc.,2002,124(46):13674-13675
27. [27]
  Mendonsa S D, Bowser M T. Anal. Chem.,2004,76(18): 5387-5392
28. [28]
  Mendonsa S D, Bowser M T. J. Am. Chem. Soc.,2005,127(26): 9382-9383
29. [29]
  Mosing R K, Mendonsa S D, Bowser M T. Anal. Chem.,2005,77(19): 6107-6112
30. [30]
  Wang J, Gao Z P, Qin S, Liu C B, Zou L L. Exp. Ther. Med.,2017,14(4): 3399-3406
31. [31]
  Qin S, Gao Z, Liu Y, Liu C, Wang J, Zou L L. Oncol. Lett.,2017,15(2): 1630-1638
32. [32]
  Beebe S J, Lassiter B P, Guo S. Cancers,2018,10(4): 97
33. [33]
  Erić A, Juranić Z, Milovanović Z, Marković I, Inić M, Stanojević-Bakić N, Vojinović-Golubović V. Pathol. Oncol. Res.,2009,15(1): 89-90
34. [34]
  Erić-Nikolić A, Milovanović Z, Sánchez D, Pekáriková A, Džodić R, Matić I Z,Tućková L, Jevrić M, Buta M, Rašković S, Juranić Z. Oncology,2012,82(1): 48-55

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乳腺癌生物标志物钙网蛋白的核酸适配体筛选及血清检测和乳腺癌细胞识别

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Screening of Aptamer for Breast Cancer Biomarker Calreticulin and Its Application to Detection of Serum and Recognition of Breast Cancer Cell

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