폐암 조기 진단을 위한 단백질 바이오마커 측정용 전압-전류법 기반의 나노바이오 분석법 개발

리징징; 스윈페이; 누드듀돈타뉴; 이혜진; Jingjing Li; Yunpei Si; Dieudonne Tanue Nde; Hye Jin Lee

Applied Chemistry for Engineering, Vol.32, No.4, 461-466, August, 2021

DOI10.14478/ace.2021.1057 Export Citation

폐암 조기 진단을 위한 단백질 바이오마커 측정용 전압-전류법 기반의 나노바이오 분석법 개발

Development of Voltammetric Nanobio-incorporated Analytical Method for Protein Biomarker Specific to Early Diagnosis of Lung Cancer

리징징, 스윈페이, 누드듀돈타뉴, 이혜진^†

경북대학교 자연과학대학 화학과

Jingjing Li, Yunpei Si, Dieudonne Tanue Nde, and Hye Jin Lee^†

Department of Chemistry, Kyungpook National University, Daegu-city 41566, Republic of Korea

E-mail:

초록

본 논문에서는 이동성이 좋고 경제적이며, 간편하게 일회용 진단칩으로 제작 가능한 스크린 프린팅 한 탄소칩 전극 [screen printed carbon electrode (SPCE)] 기반의 전압전류법 나노물질 융합형 바이오센서를 제작하여 폐암 조기진단에 활용 가능한 단백질 표지 인자 중에 하나인 heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) 단백질의 농도를 정량 분석하고자 하였다. 먼저 SPCE 표면에 금 나노입자를 전기적으로 증착한 후 크로스링커를 이용하여 hnRNP A1 에 특이적으로 결합할 수 있는 바이오리셉터인 DNA 압타머를 고정하였다. Ethanolamine을 블로킹 시약으로 사용하여 압타머와 함께 센서 표면에 고정하여 그 표면을 처리함으로써 비특이적인 생물질의 흡착에 의한 방해 신호를 최소화 하고자 하였다. DNA칩과 hnRNP A1 용액을 접촉하여 DNA와 hnRNP A1을 결합시킨 후 alkaline phosphatase (ALP) 효소로 접합한 hnRNP A1 항체(anti-hnRNP A1)을 센서칩 표면으로 주입하여 샌드위치 복합체를 형성하고, 이를 기질인 4-aminophenyl phosphate (APP)와 효소-기질 특이적 산화 반응에 의한 전류 변화를 순환 전압전류법과 시차 펄스전압전류법으로 측정하여 단백질의 농도를 정량적으로 분석하였다. 상기 산화 반응에 의한 피크 전류 변화는 순환 전압전류법과 시차 펄스 전압전류법을 사용할 때 .0.05와 -0.17 V (vs. Ag/AgCl) 전위 값에서 각각 일어났다. 개발한 나노바이오센서를 실제 정상인 혈청 시료 분석에 적용 가능함을 보여줌으로써 혈청 한 방울로 폐암의 조기진단 가능성을 제시하고자 하였다.

In this article, a portable and cost-effective voltammetric biosensor with nanoparticles was developed for the measurements of heterogeneous nuclear ribonucleoprotein A1 protein (hnRNP A1) biomarker which can potentially be used for lung cancer diagnosis. Gold nanoparticles were first electrodeposited onto screen printed carbon electrode (SPCE) followed by immobilizing a single stranded DNA aptamer specific to hnRNP A1 onto the electrode surface. Ethanolamine was also used when immobilizing DNA aptamer on the surface to prevent signals from non-specific adsorption events. Sequential injection of hnRNP A1 biomarker and anti-hnRNP A1 conjugated with alkaline phosphatase (ALP) onto the aptamer chip surface allows to form the sandwich complex of DNA aptamer/hnRNP A1/ALP-anti-hnRNP A1 on the electrode surface which further reacted with 4-aminophenyl phosphate (APP). The electrocatalytic reaction of the enzyme, ALP, and the substrate, APP, resulting in the oxidative current response changes at -0.05 and -0.17 V (vs. Ag/AgCl) against the hnRNP A1 concentration was measured using cyclic and differential pulse voltammetry, respectively. The Au nanoparticles-integrated voltammetric biosensor was applied to analyze human normal serum solutions possibly suggesting potential applicability for lung cancer diagnosis.

Keywords:Lung cancer;Early diagnosis;Protein biomarker;Voltammetric biosensor;Screen printed carbon electrode

[References]

Ferlay J, Colombet M, Soerjomataram I, Parkin DM, Pineros M, Znaor A, Bray F, Int. J. Cancer, 149, 778 (2021)
Sung H, Ferlay J, Siegel RL, Laversanne M, Soerjomataram I, Jemal A, Bray F, CA Cancer J. Clin., 71, 209 (2021)
Youn B, Trikalinos NA, Mor V, Wilson IB, Dahabreh IJ, Cancer, 126, 978 (2020)
Hanash SM, Ostrin EJ, Fahrmann JF, Transl. Lung Cancer Res., 7, 327 (2018)
Ryu HG, Jung Y, Lee N, Seo JY, Kim SW, Lee KH, Kim DY, Kim KT, Int. J. Mol. Sci., 22, 5506 (2021)
Lee SH, Goh ES, Lee HJ, Appl. Chem. Eng., 29(6), 645 (2018)
Fajri A, Goh ES, Lee SH, Lee HJ, Appl. Chem. Eng., 30(4), 429 (2019)
Hasanzadeh M, Shadjou N, Microchim. Acta, 184, 389 (2017)
Zhi LJ, Sun AL, Anal. Chim. Acta, 1134, 106 (2020)
Filik H, Avan AA, Talanta, 211, 120758 (2020)
Chiu NF, Yang HT, Front. Bioeng. Biotechnol., 8, 234 (2020)
Ranganathan V, Srinivasan S, Singh A, DeRosa MC, Anal. Biochem., 588, 113471 (2020)
Lee SY, Lee HJ, Appl. Chem. Eng., 31(6), 585 (2020)
Khanmohammadi A, Aghaie A, Vahedi E, Qazvini A, Ghanei M, Afkhami A, Hajian A, Bagheri H, Talanta, 206, 120251 (2020)
Si Y, Lee HJ, Curr. Opin. Electrochem., 22, 234 (2020)
Yang G, Xiao Z, Tang C, Deng Y, Huang H, He Z, Biosens. Bioelectron., 141, 111416 (2019)
Kava AA, Henry CS, Talanta, 221, 121553 (2021)
Roointan A, Mir TA, Wani SI, Ur RM, et al., J. Pharm. Biomed. Anal., 164, 93 (2019)
Lee S, Hayati S, Kim S, Lee HJ, Catal. Today, 359, 76 (2021)
Chen M, Wu D, Tu S, Yang C, Chen D, Xu Y, Sci. Rep., 11, 3666 (2021)
Park WC, Kim HR, Kang DB, Ryu JS, Choi KH, et al., BMC Cancer, 16, 358 (2016)
Otsuka K, Yamamoto Y, Ochiya T, Oncotarget, 9, 24718 (2018)
Zhou ZJ, Dai Z, Zhou SL, Fu XT, Zhao YM, Shi YH, Zhou J, Fan J, Int. J. Cancer, 132, 1080 (2013)
Lee SH, Park YE, Lee JE, Lee HJ, Biosens. Bioelectron., 154, 112065 (2020)
Guo R, Li Y, Ning J, Sun D, Lin L, Liu X, PloS one, 8, e62729 (2013)
Liu X, Zhou Y, Lou Y, Zhong H, Gene, 576, 791 (2016)
Zhu G, Lee HJ, Biosens. Bioelectron., 89, 959 (2017)
Chiang HC, Wang Y, Zhang Q, Levon K, Biosensors, 9, 50 (2019)
Diba FS, Kim S, Lee HJ, Biosens. Bioelectron., 72, 355 (2015)
de Oliveira O, Ferreira L, Marystela G, de Lima Leite F, Da Roz AL, Nanoscience and its Applications, William Andrew, (2016).
Leeman M, Choi J, Hansson S, Storm MU, Nilsson L, Anal. Bioanal. Chem., 410, 4867 (2018)
Ma YL, Peng JY, Zhang P, Huang L, Liu WJ, Shen TY, Chen HQ, Zhou YK, Zhang M, Chu ZX, J. Proteome Res., 8, 4525 (2009)

[Referenced By]

[1] Ferlay J, Colombet M, Soerjomataram I, Parkin DM, Pineros M, Znaor A, Bray F, Int. J. Cancer, 149, 778 (2021)

[2] Sung H, Ferlay J, Siegel RL, Laversanne M, Soerjomataram I, Jemal A, Bray F, CA Cancer J. Clin., 71, 209 (2021)

[3] Youn B, Trikalinos NA, Mor V, Wilson IB, Dahabreh IJ, Cancer, 126, 978 (2020)

[4] Hanash SM, Ostrin EJ, Fahrmann JF, Transl. Lung Cancer Res., 7, 327 (2018)

[5] Ryu HG, Jung Y, Lee N, Seo JY, Kim SW, Lee KH, Kim DY, Kim KT, Int. J. Mol. Sci., 22, 5506 (2021)

[6] Lee SH, Goh ES, Lee HJ, Appl. Chem. Eng., 29(6), 645 (2018)

[7] Fajri A, Goh ES, Lee SH, Lee HJ, Appl. Chem. Eng., 30(4), 429 (2019)

[8] Hasanzadeh M, Shadjou N, Microchim. Acta, 184, 389 (2017)

[9] Zhi LJ, Sun AL, Anal. Chim. Acta, 1134, 106 (2020)

[10] Filik H, Avan AA, Talanta, 211, 120758 (2020)

[11] Chiu NF, Yang HT, Front. Bioeng. Biotechnol., 8, 234 (2020)

[12] Ranganathan V, Srinivasan S, Singh A, DeRosa MC, Anal. Biochem., 588, 113471 (2020)

[13] Lee SY, Lee HJ, Appl. Chem. Eng., 31(6), 585 (2020)

[14] Khanmohammadi A, Aghaie A, Vahedi E, Qazvini A, Ghanei M, Afkhami A, Hajian A, Bagheri H, Talanta, 206, 120251 (2020)

[15] Si Y, Lee HJ, Curr. Opin. Electrochem., 22, 234 (2020)

[16] Yang G, Xiao Z, Tang C, Deng Y, Huang H, He Z, Biosens. Bioelectron., 141, 111416 (2019)

[17] Kava AA, Henry CS, Talanta, 221, 121553 (2021)

[18] Roointan A, Mir TA, Wani SI, Ur RM, et al., J. Pharm. Biomed. Anal., 164, 93 (2019)

[19] Lee S, Hayati S, Kim S, Lee HJ, Catal. Today, 359, 76 (2021)

[20] Chen M, Wu D, Tu S, Yang C, Chen D, Xu Y, Sci. Rep., 11, 3666 (2021)

[21] Park WC, Kim HR, Kang DB, Ryu JS, Choi KH, et al., BMC Cancer, 16, 358 (2016)

[22] Otsuka K, Yamamoto Y, Ochiya T, Oncotarget, 9, 24718 (2018)

[23] Zhou ZJ, Dai Z, Zhou SL, Fu XT, Zhao YM, Shi YH, Zhou J, Fan J, Int. J. Cancer, 132, 1080 (2013)

[24] Lee SH, Park YE, Lee JE, Lee HJ, Biosens. Bioelectron., 154, 112065 (2020)

[25] Guo R, Li Y, Ning J, Sun D, Lin L, Liu X, PloS one, 8, e62729 (2013)

[26] Liu X, Zhou Y, Lou Y, Zhong H, Gene, 576, 791 (2016)

[27] Zhu G, Lee HJ, Biosens. Bioelectron., 89, 959 (2017)

[28] Chiang HC, Wang Y, Zhang Q, Levon K, Biosensors, 9, 50 (2019)

[29] Diba FS, Kim S, Lee HJ, Biosens. Bioelectron., 72, 355 (2015)

[30] de Oliveira O, Ferreira L, Marystela G, de Lima Leite F, Da Roz AL, Nanoscience and its Applications, William Andrew, (2016).

[31] Leeman M, Choi J, Hansson S, Storm MU, Nilsson L, Anal. Bioanal. Chem., 410, 4867 (2018)

[32] Ma YL, Peng JY, Zhang P, Huang L, Liu WJ, Shen TY, Chen HQ, Zhou YK, Zhang M, Chu ZX, J. Proteome Res., 8, 4525 (2009)