title: Evaluation of antibody immobilization methods for piezoelectric biosensor application.
abstract: The immobilization of anti-Salmonella antibodies by two methods were studied and evaluated for their potential use in a piezoelectric biosensor. The optimum temperature-time combinations for the highest immobilization yields were determined for both methods. Protein A binding was found to be 67.4+/-3.8% on the gold surface which then allowed an immobilization of 42.1+/-2.09% antibody. The degree of antibody immobilization via surface aldehyde groups of glutaraldehyde (GA) on a precoated quartz crystal with polyethylenimine (PEI) was 31.6+/-0.3%. A piezoelectric probe was designed and used in dry assays to observe the frequency change due to addition of mass by the immobilization layers. The frequency changes recorded showed a better reproducibility and less added mass for the Protein A method. The frequency decrease due to microg of added antibodies was compared to frequency decrease calculated by the Sauerbrey equation. The experimental data was found to be only approximately 8% of theoretical data. The functionality of the immobilized antibodies with the Protein A method was tested with S. typhimurium in a wet chamber and the frequency decrease was compared to results of a similar system activated with PEI-GA immobilization. The frequency decreases with S. typhimurium concentration of approximately 1.5 x 10(9) CFU/ml were 50+/-2 Hz and 44+/-3 Hz for the Protein A method and PEI-GA method, respectively. It was concluded that although both methods resulted in comparable activities in terms of % immobilized protein and frequency decreases due to Salmonella binding, the Protein A method was favorable due to stability and better reproducibility of the immobilization layers.
authors.affiliation: Fiber Optic and Biosensor Research Group, Sensor and Surface Technology Partnership, FSN Research Centre, University of Rhode Island, West Kingston 2892, USA.
authors.forename: S
authors.initials: S
authors.lastname: Babacan
authors.forename: P
authors.initials: P
authors.lastname: Pivarnik
authors.forename: S
authors.initials: S
authors.lastname: Letcher
authors.forename: A G
authors.initials: AG
authors.lastname: Rand
mesh_terms: D000907:Antibodies, Bacterial / Q000276:immunology*; D015374:Biosensing Techniques; D011791:Quartz; D012475:Salmonella / Q000276:immunology* / Q000302:isolation & purification; D013205:Staphylococcal Protein A
chemical_list: D000907:Antibodies, Bacterial; D013205:Staphylococcal Protein A; D011791:Quartz