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SECM Feedback Imaging of Enzymatic Activity on Agglomerated Microbeads
Author(s) -
Wittstock Gunther,
Wilhelm Thomas,
Bahrs Sönke,
Steinrücke Peter
Publication year - 2001
Publication title -
electroanalysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.574
H-Index - 128
eISSN - 1521-4109
pISSN - 1040-0397
DOI - 10.1002/1521-4109(200105)13:8/9<669::aid-elan669>3.0.co;2-s
Subject(s) - microelectrode , materials science , agglomerate , scanning electrochemical microscopy , bead , nanotechnology , glucose oxidase , biomedical engineering , electrode , chemistry , electrochemistry , biosensor , composite material , medicine
Scanning electrochemical microscopy (SECM) was used to explore suitable conditions for feedback imaging of enzymes immobilized onto magnetic microbeads. The beads are held at the surface by a permanent magnet placed underneath the specimen. When imaging agglomerates of nitrate reductase‐covered beads it is mandatory to use an as small as possible working distance and optimized imaging conditions. The small working distance poses the risk that the microelectrode wipes away the protruding bead agglomerates. Prompted by the observation of this phenomenon, a procedure was developed that intentionally manipulates the position of microbeads on surfaces by using the microelectrode as a spatula. Lateral dragging of particles on slightly tilted specimen surfaces leads to redeposition of the beads in line‐shaped agglomerates at the point where the microelectrode‐specimen distance exceeds the diameter of the beads. This principle can be used for site‐directed immobilization and pattern formation with biochemical functionalities and is demonstrated using glucose oxidase‐modified microbeads. While dragging of microbeads can be used intentionally, it poses a considerable difficulty in analyzing the activity on small numbers of particles. In order to distinguish microbeads that stayed in place during imaging from those that were dragged by the microelectrode or from other artifacts, recording of corresponding forward and reverse scans is recommended. The subtractive combination of forward and reverse scans has values significantly different from zero only at points where beads left their place or arrived at a new position. The additive combination highlights signals from beads which stayed in the same place over the entire experiment. This procedure greatly assists in the detection of low enzymatic activity immobilized on mechanically labile samples. It should be equally applicable for analysis of metabolic activity of immobilized cells protruding from support surfaces.