Low-interference washing-free electrochemical immunosensor using glycerol-3-phosphate dehydrogenase as an enzyme label.
In washing-free electrochemical detection, various redox and reactive species cause significant interference. To minimize this interference, we report a washing-free electrochemical immunosensor using flavin adenine dinucleotide (FAD)-dependent glycerol-3-phosphate dehydrogenase (GPDH) and glycerol-3-phosphate (GP) as an enzyme label and its substrate, respectively, because the reaction of FAD-dependent dehydrogenases with dissolved O2 is slow and the level of GP preexisting in blood is low (<0.1 mM). A combination of a low electrocatalytic indium-tin oxide (ITO) electrode and fast electron-mediating Ru(NH3)6(3+) is employed to obtain a high signal-to-background ratio via proximity-dependent electron mediation of Ru(NH3)6(3+) between the ITO electrode and the GPDH label. Electrochemical oxidation of GPDH-generated Ru(NH3)6(2+) is performed at 0.05 V vs Ag/AgCl, at which point the electrochemical interference is very low. When a washing-free immunosensor is applied to cardiac troponin I detection in human serum, the calculated detection limit is approximately 10 pg/mL, indicating that the immunosensor is very sensitive in spite of the use of washing-free detection with a short detection period (10 min for incubation and 100 s for electrochemical measurement). The low-interference washing-free electrochemical immunosensor shows good promise for fast and simple point-of-care testing.
†Department of Chemistry and Chemistry Institute for Functional Materials, Pusan National University, Busan 609-735, Korea.
Gorachand
G
Dutta
†Department of Chemistry and Chemistry Institute for Functional Materials, Pusan National University, Busan 609-735, Korea.
Seonhwa
S
Park
†Department of Chemistry and Chemistry Institute for Functional Materials, Pusan National University, Busan 609-735, Korea.
Amardeep
A
Singh
†Department of Chemistry and Chemistry Institute for Functional Materials, Pusan National University, Busan 609-735, Korea.
Jeongwook
J
Seo
‡Department of Laboratory Medicine, Yonsei University College of Medicine, Seoul 135-720, Korea.
Sinyoung
S
Kim
†Department of Chemistry and Chemistry Institute for Functional Materials, Pusan National University, Busan 609-735, Korea.
Haesik
H
Yang
D055910:Antibodies, Immobilized / Q000737:chemistry; D015374:Biosensing Techniques / Q000379:methods; D055664:Electrochemical Techniques / Q000379:methods*; D004566:Electrodes; D005993:Glycerolphosphate Dehydrogenase; D006801:Humans; D007124:Immunoenzyme Techniques / Q000379:methods*; D057230:Limit of Detection; D017971:Tin Compounds / Q000737:chemistry; D019210:Troponin I / Q000032:analysis / Q000097:blood*
D055910:Antibodies, Immobilized; D017971:Tin Compounds; D019210:Troponin I; C109984:indium tin oxide; D005993:Glycerolphosphate Dehydrogenase