Dithiobissuccinimidyl propionate as an anchor for assembling peroxidases at electrodes surfaces and its application in a H2O2 biosensor.
Exposure of gold surfaces to solutions of dithiobis N-succinimidyl propionate (DTSP) gives rise to the modification of the surface with N-succinimidyl-3-thiopropionate (NSTP) which can, in turn, react with amino groups allowing for the covalent immobilization of enzymes such as horseradish peroxidase (HRP). The coverage of NSTP has been estimated to be of the order of 1.3 x 10(-10) from the charge consumed during its reductive desorption. The binding reaction of HRP with NSTP modified gold surfaces has been studied with the quartz crystal microbalance, and the results suggest that the immobilization process involves two steps in which the first (faster) appears to correspond to the rapid incorporation of the enzyme whereas the second is likely due to the slow incorporation of additional enzyme and/or reorganization of the immobilized layer. Spectrophotometric and electrochemical assays indicate that the immobilized HRP retains its enzymatic activity after immobilization onto the DTSP modified gold surface. The amount of immobilized (and active) HRP was estimated from QCM and spectrophotometric measurements to be of the order of 1.5 x 10(-11) mol/cm2. A peroxide biosensor was developed making use of a gold surface modified with DTSP and HRP employing Os and Ru complexes of 1,10-phenanthroline 5,6-dione (phen-dione) of the type [M(phendione)x(L)3-x]+2 (where L = 1,10-phenanthroline or 2,2'-bipyridine, x = 1-3) as mediators with the quinone moieties being the active component. The efficiency of the mediators increased with increasing number of phendione ligands.
Darder
M
M
Department of Chemistry and Chemical Biology, Baker Laboratory, Cornell University, Ithaca, New York 14853-1301, USA.
Takada
K
K
Pariente
F
F
Lorenzo
E
E
Abruña
H D
HD
D000327:Adsorption; D015374:Biosensing Techniques; D002384:Catalysis; D003432:Cross-Linking Reagents / Q000737:chemistry*; D004566:Electrodes; D006418:Heme / Q000032:analysis; D006861:Hydrogen Peroxide / Q000032:analysis*; D010084:Oxidation-Reduction; D010544:Peroxidases / Q000737:chemistry*; D013388:Succinimides / Q000737:chemistry*
D003432:Cross-Linking Reagents; D013388:Succinimides; D006418:Heme; D006861:Hydrogen Peroxide; D010544:Peroxidases; C011240:dithiobis(succinimidylpropionate)