Label-free Electrochemical Immunosensor for Ultrasensitive Detection of KLK4 Using Conjugated Polymer Modified Sensing Platform

Elif Burcu Aydın

doi:10.15671/hjbc.1633456

Research Article

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Year 2025, Volume: 53 Issue: 3, 43 - 55, 01.07.2025

Elif Burcu Aydın

https://doi.org/10.15671/hjbc.1633456

Abstract

References

S. Singh, A.A. Gill, M. Nlooto, R. Karpoormath, Prostate cancer biomarkers detection using nanoparticles based electrochemical biosensors, Biosens. Bioelectron. 137 (2019) 213-221.
A. Moradi, S. Srinivasan, J. Clements, J. Batra, Beyond the biomarker role: prostate-specific antigen (PSA) in the prostate cancer microenvironment, Cancer Metastasis Rev. 38 (2019) 333-346.
S.O. Oseni, C. Naar, M. Pavlović, W. Asghar, J.X. Hartmann, G.B. Fields, N. Esiobu, J. Kumi-Diaka, The molecular basis and clinical consequences of chronic inflammation in prostatic diseases: prostatitis, benign prostatic hyperplasia, and prostate Cancer, Cancers 15(12) (2023) 3110.
S.K. Hong, Kallikreins as biomarkers for prostate cancer, BioMed research international 2014(1) (2014) 526341.
M. Avgeris, K. Stravodimos, A. Scorilas, Kallikrein‐related peptidase 4 gene (KLK4) in prostate tumors: Quantitative expression analysis and evaluation of its clinical significance, The Prostate 71(16) (2011) 1780-1789.
C.H. Day, G.R. Fanger, M.W. Retter, B.L. Hylander, R.B. Penetrante, R.L. Houghton, X. Zhang, P.D. McNeill, M. Nolasco, R. Badaro, Characterization of KLK4 expression and detection of KLK4-specific antibody in prostate cancer patient sera, Oncogene 21(46) (2002) 7114-7120.
T. Kryza, M.L. Silva, D. Loessner, N. Heuzé-Vourc'h, J. Clements, The kallikrein-related peptidase family: Dysregulation and functions during cancer progression, Biochimie 122 (2016) 283-299.
L. Wang, T. Skotland, V. Berge, K. Sandvig, A. Llorente, Exosomal proteins as prostate cancer biomarkers in urine: from mass spectrometry discovery to immunoassay-based validation, Eur. J. Pharm. Sci. 98 (2017) 80-85.
K. Fujita, N. Nonomura, Urinary biomarkers of prostate cancer, Int. J. Urol. 25(9) (2018) 770-779.
J. Korram, A.C. Anbalagan, A. Banerjee, S.N. Sawant, Bio-conjugated carbon dots for the bimodal detection of prostate cancer biomarkers via sandwich fluorescence and electrochemical immunoassays, J. Mater. Chem. B 12(3) (2024) 742-751.
Y. Hu, S. Lv, J. Wan, C. Zheng, D. Shao, H. Wang, Y. Tao, M. Li, Y. Luo, Recent advances in nanomaterials for prostate cancer detection and diagnosis, J. Mater. Chem. B 10(26) (2022) 4907-4934.
A.C. Murphy, M.E. Wechsler, N.A. Peppas, Recent advancements in biosensing approaches for screening and diagnostic applications, Curr. Opin. Biomed. Eng. 19 (2021) 100318.
W. Wen, X. Yan, C. Zhu, D. Du, Y. Lin, Recent advances in electrochemical immunosensors, Anal. Chem. 89(1) (2017) 138-156.
C. Robinson, V.B. Juska, A. O'Riordan, Surface chemistry applications and development of immunosensors using electrochemical impedance spectroscopy: A comprehensive review, Environ. Res. (2023) 116877.
M. Aydın, E.B. Aydın, M.K. Sezgintürk, Functionalized magnetic nanoparticles for electrochemical magneto biosensing of PSMA cancer biomarker, New J. Chem. 48(13) (2024) 5769-5781.
E.B. Aydın, M. Aydın, M.K. Sezgintürk, Fabrication of a highly sensitive conductive copolymer layer modified immunosensing tool for cytokeratin-19 fragment detection, Microchem. J. 206 (2024) 111464.
Y. Wan, Y. Su, X. Zhu, G. Liu, C. Fan, Development of electrochemical immunosensors towards point of care diagnostics, Biosens. Bioelectron. 47 (2013) 1-11.
H.S. Magar, R.Y. Hassan, A. Mulchandani, Electrochemical impedance spectroscopy (EIS): Principles, construction, and biosensing applications, Sensors 21(19) (2021) 6578.
M. Aydın, E.B. Aydın, M.K. Sezgintürk, Ultrasensitive detection of NSE employing a novel electrochemical immunosensor based on a conjugated copolymer, Analyst 149(5) (2024) 1632-1644.
E.B. Aydın, M. Aydın, M.K. Sezgintürk, Electrochemical immunosensor for detection of CCR4 cancer biomarker in human serum: An alternative strategy for modification of disposable ITO electrode, Macromol. Biosci. 21(2) (2021) 2000267.
A. Chakraborty, P. Dutta, A. Wakankar, C. RoyChaudhuri, Recent field effect transistors and electrical impedance spectroscopy based biosensing strategies for cancer biomarker screening: A mini review, Biosens. Bioelectron.:X 12 (2022) 100253.
R. Cancelliere, E. Paialunga, A. Grattagliano, L. Micheli, Label-free electrochemical immunosensors: a practical guide, TrAC, Trends Anal. Chem. (2024) 117949.
L. Zhang, W. Du, A. Nautiyal, Z. Liu, X. Zhang, Recent progress on nanostructured conducting polymers and composites: synthesis, application and future aspects, Sci. China. Mater. 61(3) (2018) 303-352.
T. Nezakati, A. Seifalian, A. Tan, A.M. Seifalian, Conductive polymers: opportunities and challenges in biomedical applications, Chem. Rev. 118(14) (2018) 6766-6843.
G. Anantha-Iyengar, K. Shanmugasundaram, M. Nallal, K.-P. Lee, M.J. Whitcombe, D. Lakshmi, G. Sai-Anand, Functionalized conjugated polymers for sensing and molecular imprinting applications, Prog. Polym. Sci. 88 (2019) 1-129.
M. Aydın, A sensitive and selective approach for detection of IL 1α cancer biomarker using disposable ITO electrode modified with epoxy-substituted polythiophene polymer, Biosens. Bioelectron. 144 (2019) 111675.
P.F. Cañamero, J.L. de la Fuente, E.L. Madruga, M. Fernández‐García, Atom transfer radical polymerization of glycidyl methacrylate: a functional monomer, Macromol. Chem. Phy. 205(16) (2004) 2221-2228.
M. Jonsson, D. Nyström, O. Nordin, E. Malmström, Surface modification of thermally expandable microspheres by grafting poly (glycidyl methacrylate) using ARGET ATRP, Eur. Pol. J. 45(8) (2009) 2374-2382.
E. Sahin, P. Camurlu, L. Toppare, V.M. Mercore, I. Cianga, Y. Yagˇcı, Conducting copolymers of thiophene functionalized polystyrenes with thiophene, J. Electroanal. Chem. 579(2) (2005) 189-197.
M. Wang, P. Baek, L. Voorhaar, E.W.C. Chan, A. Nelson, D. Barker, J. Travas-Sejdic, Long side-chain grafting imparts intrinsic adhesiveness to poly (thiophene phenylene) conjugated polymer, Eur. Pol. J. 109 (2018) 237-247.
J. Muñoz, R. Montes, M. Baeza, Trends in electrochemical impedance spectroscopy involving nanocomposite transducers: Characterization, architecture surface and bio-sensing, TrAC, Trends Anal. Chem. 97 (2017) 201-215.
E.P. Randviir, C.E. Banks, Electrochemical impedance spectroscopy: an overview of bioanalytical applications, Anal. Meth. 5(5) (2013) 1098-1115.
Rajesh, S. Singal, R.K. Kotnala, Single frequency impedance analysis on reduced graphene oxide screen-printed electrode for biomolecular detection, Appl. Biochem. Biotechnol. 183 (2017) 672-683.
E.B. Aydın, M. Aydın, M.K. Sezgintürk, Fabrication of electrochemical immunosensor based on acid-substituted poly (pyrrole) polymer modified disposable ITO electrode for sensitive detection of CCR4 cancer biomarker in human serum, Talanta 222 (2021) 121487.
E.B. Aydın, M. Aydın, A. Yuzer, M. Ince, K. Ocakoglu and M. K. Sezginturk, Detection of Kallikrein-related Peptidase 4 with a label-free electrochemical impedance biosensor based on a zinc (II) phthalocyanine tetracarboxylic acid-functionalized disposable indium tin oxide electrode, ACS Biomater. Sci. Eng. 7(3) (2021) 1192-1201.
E.B. Aydın, M. Aydın, M. K. Sezginturk, Construction of succinimide group substituted polythiophene polymer functionalized sensing platform for ultrasensitive detection of KLK 4 cancer biomarker, Sens. Actuators, B, 325 (2020) 128788.

Label-free Electrochemical Immunosensor for Ultrasensitive Detection of KLK4 Using Conjugated Polymer Modified Sensing Platform

Year 2025, Volume: 53 Issue: 3, 43 - 55, 01.07.2025

Elif Burcu Aydın

https://doi.org/10.15671/hjbc.1633456

Abstract

In the present research, a label-free impedimetric kallikrein 4 (KLK4) biosensor was fabricated by using a low-cost and disposable indium tin oxide (ITO) electrode and utilized for KLK4 detection in human serum. The ITO electrode was modified first by spin-coating of epoxy-substituted poly(thiophene) (Poly(TEpx)) conjugated polymer and thence anti-KLK4 antibody immobilization via the epoxy ends on the polymer. The specific interaction between the anti-KLK4 antibody present on the working electrode surface and the KLK4 antigen caused increases in the impedimetric response, and the KLK4 antigen was determined in a linear concentration range from 0.025 to 12.5 pg/mL with a detection limit of 7.78 fg/mL. Furthermore, the developed biosensor had acceptable repeatability and reproducibility, long storage stability, and high selectivity to KLK4 antigen. The applicability of the immunosensor was tested in human serum samples and satisfactory results were obtained.

Keywords

Kallikrein 4, electrochemical impedance spectroscopy, prostate cancer, conjugated polymer.

References

S. Singh, A.A. Gill, M. Nlooto, R. Karpoormath, Prostate cancer biomarkers detection using nanoparticles based electrochemical biosensors, Biosens. Bioelectron. 137 (2019) 213-221.
A. Moradi, S. Srinivasan, J. Clements, J. Batra, Beyond the biomarker role: prostate-specific antigen (PSA) in the prostate cancer microenvironment, Cancer Metastasis Rev. 38 (2019) 333-346.
S.O. Oseni, C. Naar, M. Pavlović, W. Asghar, J.X. Hartmann, G.B. Fields, N. Esiobu, J. Kumi-Diaka, The molecular basis and clinical consequences of chronic inflammation in prostatic diseases: prostatitis, benign prostatic hyperplasia, and prostate Cancer, Cancers 15(12) (2023) 3110.
S.K. Hong, Kallikreins as biomarkers for prostate cancer, BioMed research international 2014(1) (2014) 526341.
M. Avgeris, K. Stravodimos, A. Scorilas, Kallikrein‐related peptidase 4 gene (KLK4) in prostate tumors: Quantitative expression analysis and evaluation of its clinical significance, The Prostate 71(16) (2011) 1780-1789.
C.H. Day, G.R. Fanger, M.W. Retter, B.L. Hylander, R.B. Penetrante, R.L. Houghton, X. Zhang, P.D. McNeill, M. Nolasco, R. Badaro, Characterization of KLK4 expression and detection of KLK4-specific antibody in prostate cancer patient sera, Oncogene 21(46) (2002) 7114-7120.
T. Kryza, M.L. Silva, D. Loessner, N. Heuzé-Vourc'h, J. Clements, The kallikrein-related peptidase family: Dysregulation and functions during cancer progression, Biochimie 122 (2016) 283-299.
L. Wang, T. Skotland, V. Berge, K. Sandvig, A. Llorente, Exosomal proteins as prostate cancer biomarkers in urine: from mass spectrometry discovery to immunoassay-based validation, Eur. J. Pharm. Sci. 98 (2017) 80-85.
K. Fujita, N. Nonomura, Urinary biomarkers of prostate cancer, Int. J. Urol. 25(9) (2018) 770-779.
J. Korram, A.C. Anbalagan, A. Banerjee, S.N. Sawant, Bio-conjugated carbon dots for the bimodal detection of prostate cancer biomarkers via sandwich fluorescence and electrochemical immunoassays, J. Mater. Chem. B 12(3) (2024) 742-751.
Y. Hu, S. Lv, J. Wan, C. Zheng, D. Shao, H. Wang, Y. Tao, M. Li, Y. Luo, Recent advances in nanomaterials for prostate cancer detection and diagnosis, J. Mater. Chem. B 10(26) (2022) 4907-4934.
A.C. Murphy, M.E. Wechsler, N.A. Peppas, Recent advancements in biosensing approaches for screening and diagnostic applications, Curr. Opin. Biomed. Eng. 19 (2021) 100318.
W. Wen, X. Yan, C. Zhu, D. Du, Y. Lin, Recent advances in electrochemical immunosensors, Anal. Chem. 89(1) (2017) 138-156.
C. Robinson, V.B. Juska, A. O'Riordan, Surface chemistry applications and development of immunosensors using electrochemical impedance spectroscopy: A comprehensive review, Environ. Res. (2023) 116877.
M. Aydın, E.B. Aydın, M.K. Sezgintürk, Functionalized magnetic nanoparticles for electrochemical magneto biosensing of PSMA cancer biomarker, New J. Chem. 48(13) (2024) 5769-5781.
E.B. Aydın, M. Aydın, M.K. Sezgintürk, Fabrication of a highly sensitive conductive copolymer layer modified immunosensing tool for cytokeratin-19 fragment detection, Microchem. J. 206 (2024) 111464.
Y. Wan, Y. Su, X. Zhu, G. Liu, C. Fan, Development of electrochemical immunosensors towards point of care diagnostics, Biosens. Bioelectron. 47 (2013) 1-11.
H.S. Magar, R.Y. Hassan, A. Mulchandani, Electrochemical impedance spectroscopy (EIS): Principles, construction, and biosensing applications, Sensors 21(19) (2021) 6578.
M. Aydın, E.B. Aydın, M.K. Sezgintürk, Ultrasensitive detection of NSE employing a novel electrochemical immunosensor based on a conjugated copolymer, Analyst 149(5) (2024) 1632-1644.
E.B. Aydın, M. Aydın, M.K. Sezgintürk, Electrochemical immunosensor for detection of CCR4 cancer biomarker in human serum: An alternative strategy for modification of disposable ITO electrode, Macromol. Biosci. 21(2) (2021) 2000267.
A. Chakraborty, P. Dutta, A. Wakankar, C. RoyChaudhuri, Recent field effect transistors and electrical impedance spectroscopy based biosensing strategies for cancer biomarker screening: A mini review, Biosens. Bioelectron.:X 12 (2022) 100253.
R. Cancelliere, E. Paialunga, A. Grattagliano, L. Micheli, Label-free electrochemical immunosensors: a practical guide, TrAC, Trends Anal. Chem. (2024) 117949.
L. Zhang, W. Du, A. Nautiyal, Z. Liu, X. Zhang, Recent progress on nanostructured conducting polymers and composites: synthesis, application and future aspects, Sci. China. Mater. 61(3) (2018) 303-352.
T. Nezakati, A. Seifalian, A. Tan, A.M. Seifalian, Conductive polymers: opportunities and challenges in biomedical applications, Chem. Rev. 118(14) (2018) 6766-6843.
G. Anantha-Iyengar, K. Shanmugasundaram, M. Nallal, K.-P. Lee, M.J. Whitcombe, D. Lakshmi, G. Sai-Anand, Functionalized conjugated polymers for sensing and molecular imprinting applications, Prog. Polym. Sci. 88 (2019) 1-129.
M. Aydın, A sensitive and selective approach for detection of IL 1α cancer biomarker using disposable ITO electrode modified with epoxy-substituted polythiophene polymer, Biosens. Bioelectron. 144 (2019) 111675.
P.F. Cañamero, J.L. de la Fuente, E.L. Madruga, M. Fernández‐García, Atom transfer radical polymerization of glycidyl methacrylate: a functional monomer, Macromol. Chem. Phy. 205(16) (2004) 2221-2228.
M. Jonsson, D. Nyström, O. Nordin, E. Malmström, Surface modification of thermally expandable microspheres by grafting poly (glycidyl methacrylate) using ARGET ATRP, Eur. Pol. J. 45(8) (2009) 2374-2382.
E. Sahin, P. Camurlu, L. Toppare, V.M. Mercore, I. Cianga, Y. Yagˇcı, Conducting copolymers of thiophene functionalized polystyrenes with thiophene, J. Electroanal. Chem. 579(2) (2005) 189-197.
M. Wang, P. Baek, L. Voorhaar, E.W.C. Chan, A. Nelson, D. Barker, J. Travas-Sejdic, Long side-chain grafting imparts intrinsic adhesiveness to poly (thiophene phenylene) conjugated polymer, Eur. Pol. J. 109 (2018) 237-247.
J. Muñoz, R. Montes, M. Baeza, Trends in electrochemical impedance spectroscopy involving nanocomposite transducers: Characterization, architecture surface and bio-sensing, TrAC, Trends Anal. Chem. 97 (2017) 201-215.
E.P. Randviir, C.E. Banks, Electrochemical impedance spectroscopy: an overview of bioanalytical applications, Anal. Meth. 5(5) (2013) 1098-1115.
Rajesh, S. Singal, R.K. Kotnala, Single frequency impedance analysis on reduced graphene oxide screen-printed electrode for biomolecular detection, Appl. Biochem. Biotechnol. 183 (2017) 672-683.
E.B. Aydın, M. Aydın, M.K. Sezgintürk, Fabrication of electrochemical immunosensor based on acid-substituted poly (pyrrole) polymer modified disposable ITO electrode for sensitive detection of CCR4 cancer biomarker in human serum, Talanta 222 (2021) 121487.
E.B. Aydın, M. Aydın, A. Yuzer, M. Ince, K. Ocakoglu and M. K. Sezginturk, Detection of Kallikrein-related Peptidase 4 with a label-free electrochemical impedance biosensor based on a zinc (II) phthalocyanine tetracarboxylic acid-functionalized disposable indium tin oxide electrode, ACS Biomater. Sci. Eng. 7(3) (2021) 1192-1201.
E.B. Aydın, M. Aydın, M. K. Sezginturk, Construction of succinimide group substituted polythiophene polymer functionalized sensing platform for ultrasensitive detection of KLK 4 cancer biomarker, Sens. Actuators, B, 325 (2020) 128788.

There are 36 citations in total.

Details

Primary Language	English
Subjects	Electroanalytical Chemistry, Sensor Technology
Journal Section	Research Article
Authors	Elif Burcu Aydın 0000-0002-1982-4128
Publication Date	July 1, 2025
Submission Date	February 5, 2025
Acceptance Date	June 26, 2025
Published in Issue	Year 2025 Volume: 53 Issue: 3

Cite

APA	Aydın, E. B. (2025). Label-free Electrochemical Immunosensor for Ultrasensitive Detection of KLK4 Using Conjugated Polymer Modified Sensing Platform. Hacettepe Journal of Biology and Chemistry, 53(3), 43-55. https://doi.org/10.15671/hjbc.1633456
AMA	Aydın EB. Label-free Electrochemical Immunosensor for Ultrasensitive Detection of KLK4 Using Conjugated Polymer Modified Sensing Platform. HJBC. July 2025;53(3):43-55. doi:10.15671/hjbc.1633456
Chicago	Aydın, Elif Burcu. “Label-Free Electrochemical Immunosensor for Ultrasensitive Detection of KLK4 Using Conjugated Polymer Modified Sensing Platform”. Hacettepe Journal of Biology and Chemistry 53, no. 3 (July 2025): 43-55. https://doi.org/10.15671/hjbc.1633456.
EndNote	Aydın EB (July 1, 2025) Label-free Electrochemical Immunosensor for Ultrasensitive Detection of KLK4 Using Conjugated Polymer Modified Sensing Platform. Hacettepe Journal of Biology and Chemistry 53 3 43–55.
IEEE	E. B. Aydın, “Label-free Electrochemical Immunosensor for Ultrasensitive Detection of KLK4 Using Conjugated Polymer Modified Sensing Platform”, HJBC, vol. 53, no. 3, pp. 43–55, 2025, doi: 10.15671/hjbc.1633456.
ISNAD	Aydın, Elif Burcu. “Label-Free Electrochemical Immunosensor for Ultrasensitive Detection of KLK4 Using Conjugated Polymer Modified Sensing Platform”. Hacettepe Journal of Biology and Chemistry 53/3 (July 2025), 43-55. https://doi.org/10.15671/hjbc.1633456.
JAMA	Aydın EB. Label-free Electrochemical Immunosensor for Ultrasensitive Detection of KLK4 Using Conjugated Polymer Modified Sensing Platform. HJBC. 2025;53:43–55.
MLA	Aydın, Elif Burcu. “Label-Free Electrochemical Immunosensor for Ultrasensitive Detection of KLK4 Using Conjugated Polymer Modified Sensing Platform”. Hacettepe Journal of Biology and Chemistry, vol. 53, no. 3, 2025, pp. 43-55, doi:10.15671/hjbc.1633456.
Vancouver	Aydın EB. Label-free Electrochemical Immunosensor for Ultrasensitive Detection of KLK4 Using Conjugated Polymer Modified Sensing Platform. HJBC. 2025;53(3):43-55.

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