Preparation and evaluation of spore-specific affinity-augmented bio-imprinted beads
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A novel, affinity-augmented, bacterial spore-imprinted, bead material was synthesized, based on a procedure developed for vegetative bacteria. The imprinted beads were intended as a front-end spore capture/concentration stage of an integrated biological detection system. Our approach involved embedding bead surfaces with Bacillus thuringiensis kurstaki (Bt) spores (as a surrogate for Bacillus anthracis) during synthesis. Subsequent steps involved lithographic deactivation using a perfluoroether; spore removal to create imprint sites; and coating imprints with the lectin, concanavalin A, to provide general affinity. The synthesis of the intended material with the desired imprints was verified by scanning electron and confocal laser-scanning microscopy. The material was evaluated using spore-binding assays with either Bt or Bacillus subtilis (Bs) spores. The binding assays indicated strong spore-binding capability and a robust imprinting effect that accounted for 25% additional binding over non-imprinted controls. The binding assay results also indicated that further refinement of the surface deactivation procedure would enhance the performance of the imprinted substrate.
KeywordsBio-imprinted beads Selective spore capture/concentration Analysis of biological pathogens
We gratefully acknowledge the Laboratory Directed Research and Development program for funding this research through the Homeland Security Initiative at Pacific Northwest National Laboratory. We also acknowledge the assistance of Catherine E. Petersen in collecting MALDI spectra. Pacific Northwest National Laboratory is a multi-program national laboratory operated by the Battelle Memorial Institute for the U.S. Department of Energy under contract DE-AC06-76RLO 1830.
- 9.Skaggs R, Straub T, Wright B, Bruckner-Lea C, Harvey S (2004) In: Mays L (ed) Water supply systems security, chapt 18. McGraw Hill, New York, pp 18.1–18.21Google Scholar
- 10.Berry ED, Siragusa GR (1999) J Rapid Methods Autom Microbiol 7:7–23Google Scholar
- 12.Marshall, KC (1991) In: Mozes N, Handley PS, Busscher HJ, Rouxhet PG (eds) Microbial cell surface analysis. VCH Publishers, New York, pp 5–19Google Scholar
- 19.Perez N, Alexander C, Vulfson EN (2001) In: Sellergren B (ed) Molecularly imprinted polymers: man-made mimics of antibodies and their applications in analytical chemistry. Elsevier, Amsterdam, The Netherlands, pp 295–303Google Scholar
- 20.Cunliffe D, Alexander C (2002) In: Yan M, Ramstrom O (eds) Molecular imprinted materials: science and technology. Dekker, New York, pp 249–283Google Scholar
- 21.Alexander C, Kirsch N, Whitcombe MJ (2004) In: Davis FJ (ed) Polymer chemistry: a practical approach in chemistry. Oxford University Press, Oxford, UK, pp 201–214Google Scholar
- 22.Nicholson WL, Setlow P (1990) In: Harwood CR, Cutting SM (eds) Molecular biological methods for bacillus. Wiley, Chichester, England, pp 391–450Google Scholar
- 24.Koshikawa T, Yamazaki M, Yoshimi M, Ogawa S, Yamada A, Watabe K, Torii M (1989) J Gen Microbiol 135:271–2722Google Scholar