Biomedical Engineering Reference
In-Depth Information
an analyte, or group of analytes, to a measurable response. Thus, for electrochemical
biosensors based on direct electron transfer of protein, how to immobilize the biologi-
cal recognition elements, such as redox proteins, onto the signal transducers, here the
electrodes, are an important process.
17.2.1.1 Adsorption of protein
The physical adsorption of protein onto the surface of an electrode is a simple immobi-
lization method. The adsorption is obtained by volatilizing the buffers containing pro-
teins. The physical adsorption needs no chemical reagent, seldom activation and rinse,
so that the bioactivities of the immobilized proteins can be retained well. However,
the immobilized proteins are easy to break off from the electrode, which restrict broad
applications of this method. Below are some examples of the physical adsorption of
proteins immobilized on electrodes.
Ikariyama [2] described a unique method for the preparation of a glucose oxidase
(GOD) electrode in their work. The method is based on two electrochemical proc-
esses, i.e. electrochemical adsorption of GOD molecules and electrochemical growth
of porous electrode. GOD immobilized in the growing matrix of platinum black par-
ticles is employed for the microfabrication of the enzyme electrode. It demonstrated
high performance with high sensitivity and fast responsiveness.
Suaud-Chagny and Gonon [3] presented a new procedure for protein immobiliza-
tion adapted to carbon microelectrode characteristics. The principle of this method
of immobilization is based on the association of the protein with an inert porous fi lm
immobilized around the active tip of the electrode. For this purpose the carbon was
coated with an inert, electrochemically obtained protein sheath (bovine serum albu-
min, BSA) a few micrometers thick. Then the sheath around the fi ber was impregnated
with lactate dehydrogenase (LDH), which could be immobilized onto the electrode and
resulted in an electrode sensitive to pyruvate.
17.2.1.2 Covalent bonding of protein
Protein is immobilized by combining with the surface of the electrode through a cova-
lent bond, which is called covalent bonding of protein. The process requires low tem-
perature (0ÂșC), low ion intensity, and physiological pH conditions. Although covalent
bonding onto the surface of an electrode is more diffi cult than adsorption, it can pro-
vide a more stable immobilized protein.
Moody [4] described in his paper a miniature enzyme electrode consisting of GOD
covalently attached to a silanized and anodized platinum wire surface via the bifunc-
tional glutaraldehyde enzyme-immobilizing reagent. Its response characteristics were
determined in a three-electrode amperometric mode by monitoring the anodic decom-
position of hydrogen. The system exhibited good linearity (for glucose concentrations
of 0.1-20 mM), where log ( I /A)
0.992 log ([glucose]/M)
3.94, with a correlation
coeffi cient of 0.999. Response times (
30 s) were short, and a
lifetime of 9 h was obtained for continuous exposure to 2.5 and 10 mM glucose.
25 s) and wash times (
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