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Proceedings Paper

Integrating biotinylated polyalkylthiophene thin films with biological macromolecules: biosensing organophosphorus pesticides and metal ions with surface immobilized alkaline phosphatase utilizing chemiluminescence measurements
Author(s): Rajiv Pande; S. Kamtekar; Madhu Sudhan Rao Ayyagari; Kenneth A. Marx; Jayant Kumar; Sukant K. Tripathy; David L. Kaplan
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Paper Abstract

We describe a methodology for immobilizing the enzyme alkaline phosphatase onto a glass surface using a novel biotinylated copolymer poly (3-undecylthiophene-co-3- thiophenecarboxaldehyde) 6-biotinamido hexanohydrazide attached hydrophobically to silanized glass. The biotin-streptavidin protein interaction is used to carry out this immobilization. Alkaline phosphatase catalyzes the dephosphorylation of a class of macrocyclic compounds: including CSPD {chloro 3-[4-methoxy spiro(1,2 dioxetane-3-2-trichloro-(3.3.1.1)-decan]-4 yl}phenyl phosphate to a product species which emits energy by chemiluminescence. We can detect this chemiluminescence signal with a photomultiplier tube for both enzymatic catalysis in solution and the surface immobilized enzyme (streptavidin conjugate). This enzyme is inhibited by the organophosphorus class of pesticides as well as nerve agents. The enzyme is also inhibited by Be(II), Bi(III) as well as excess Zn(II), while the apoenzyme is reactivated by Zn(II). We demonstrate in this study that two representative organophosphorus pesticides inhibit the enzymatic production of chemiluminescent products. This is true for the enzyme conjugate both free in solution and immobilized. We can detect pesticides down to about 50 ppb for the enzyme in solution and 500 ppb for surface immobilized enzyme in a 100 (mu) l capillary. Detection of Zn(II) by apoenzyme reactivation occurs down to 3 ppb. Be(II) and Bi(III) are detected by inhibition down to 1 ppm.

Paper Details

Date Published: 22 May 1995
PDF: 11 pages
Proc. SPIE 2441, Smart Structures and Materials 1995: Smart Materials, (22 May 1995); doi: 10.1117/12.209798
Show Author Affiliations
Rajiv Pande, Univ. of Massachusetts/Lowell (United States)
S. Kamtekar, Univ. of Massachusetts/Lowell (United States)
Madhu Sudhan Rao Ayyagari, Univ. of Massachusetts/Lowell (United States)
Kenneth A. Marx, Univ. of Massachusetts/Lowell (United States)
Jayant Kumar, Univ. of Massachusetts/Lowell (United States)
Sukant K. Tripathy, Univ. of Massachusetts/Lowell (United States)
David L. Kaplan, U.S. Army Natick Research, Development, and Engineering Ctr. (United States)


Published in SPIE Proceedings Vol. 2441:
Smart Structures and Materials 1995: Smart Materials
A. Peter Jardine, Editor(s)

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