TY - JOUR
T1 - Femtomolar Detection of Thrombin in Serum and Cerebrospinal Fluid via Direct Electrocatalysis of Oxygen Reduction by the Covalent G4-Hemin-Aptamer Complex
AU - Malecka, Kamila
AU - Ferapontova, Elena E.
PY - 2021/8/18
Y1 - 2021/8/18
N2 - Thrombin, a serine protease playing a central role in the coagulation cascade reactions and a potent neurotoxin produced by injured brain endothelial cells, is a recognized cardiac biomarker and a critical biomarker for Alzheimer's disease development. Both in vivo and in vitro, its low physiological concentrations and nonspecific binding of other components of physiological fluids complicate electroanalysis of thrombin. Here, femtomolar levels of thrombin in serum and an artificial cerebrospinal fluid (CSF) were detected by the indicator-free electrochemical methodology exploiting the O2 reduction reaction directly, with no electron transfer mediators, electrocatalyzed by the covalent G4-hemin DNAzyme complex naturally self-assembling upon thrombin binding to the hemin-modified 29-mer DNA aptamer sequence tethered to gold via an alkanethiol linker. Coadsorbed PEG inhibited nonspecific protein binding and allowed the sought signal resolution. The proposed assay exploiting the "oxidase"activity of G4-hemin DNAzyme does not require any coreactants necessary for the traditional peroxidase activity-based assays with this DNAzyme, such as H2O2 and redox mediators, or solution deaeration and allows fast, overall 30 min analysis of thrombin in aerated buffer, CSF, and 1% human serum solutions. This pioneer approach exploiting the oxidase activity G4-hemin DNAzyme is simple, sensitive, and selective and represents a new tool for ultrasensitive electrocatalytic assays based on simple and efficient O2-dependent DNAzyme labels.
AB - Thrombin, a serine protease playing a central role in the coagulation cascade reactions and a potent neurotoxin produced by injured brain endothelial cells, is a recognized cardiac biomarker and a critical biomarker for Alzheimer's disease development. Both in vivo and in vitro, its low physiological concentrations and nonspecific binding of other components of physiological fluids complicate electroanalysis of thrombin. Here, femtomolar levels of thrombin in serum and an artificial cerebrospinal fluid (CSF) were detected by the indicator-free electrochemical methodology exploiting the O2 reduction reaction directly, with no electron transfer mediators, electrocatalyzed by the covalent G4-hemin DNAzyme complex naturally self-assembling upon thrombin binding to the hemin-modified 29-mer DNA aptamer sequence tethered to gold via an alkanethiol linker. Coadsorbed PEG inhibited nonspecific protein binding and allowed the sought signal resolution. The proposed assay exploiting the "oxidase"activity of G4-hemin DNAzyme does not require any coreactants necessary for the traditional peroxidase activity-based assays with this DNAzyme, such as H2O2 and redox mediators, or solution deaeration and allows fast, overall 30 min analysis of thrombin in aerated buffer, CSF, and 1% human serum solutions. This pioneer approach exploiting the oxidase activity G4-hemin DNAzyme is simple, sensitive, and selective and represents a new tool for ultrasensitive electrocatalytic assays based on simple and efficient O2-dependent DNAzyme labels.
KW - aptamer
KW - biosensor
KW - DNAzyme
KW - electrocatalysis
KW - G4-hemin covalent complex
KW - thrombin
UR - http://www.scopus.com/inward/record.url?scp=85106604887&partnerID=8YFLogxK
U2 - 10.1021/acsami.1c03784
DO - 10.1021/acsami.1c03784
M3 - Journal article
C2 - 33878266
AN - SCOPUS:85106604887
SN - 1944-8244
VL - 13
SP - 37979
EP - 37988
JO - ACS Applied Materials & Interfaces
JF - ACS Applied Materials & Interfaces
IS - 32
ER -