Optimization and characterization of a biosensor assembly for detection of Salmonella Typhimurium
The performance of biosensors depends directly on the strategies adopted during their development. In this paper, a fast and sensitive biosensor for Salmonella Typhimurium detection was assembled by using optimization studies in separate stages. The pre-treatment assays, biomolecular immobilization (primary antibody and protein A concentrations), and analytical response (hydroquinone and hydrogen peroxide concentrations) were optimized via voltammetric methods. In the biosensor assembly, a gold surface was modified via the self-assembled monolayer technique (SAM) using cysteamine thiol and protein A for immobilization of anti-Salmonella antibody. The analytical response of the biosensor was obtained through the use of a secondary antibody labeled with a peroxidase enzyme, and the signal was evaluated by applying the chronoamperometry technique. The biosensor was characterized by infrared spectroscopy and cyclic voltammetry. Optimization of protein A and primary antibody concentrations enabled higher analytical signals of 7.5 and 75 mg mL−1, respectively, to be achieved. The hydroquinone and H2O2 concentrations selected were 3 and 300 mM, respectively. The biosensor developed attained a very low detection limit of 10 CFU mL−1 and a fast response with a final detection time of 125 min. These results indicate that this biosensor is very promising for the food safety and emergency response applications.
KeywordsSalmonella Immunosensor Amperometric Pathogen Rapid detection
The authors would like to thank the Brazilian agencies, CNPq, FUNCAP, and CAPES, for their financial support, Embrapa Tropical Agroindustry and National Center of Energy and Materials Research (CNPEM). Mention of trade names or commercial products in this publication is solely for the purpose of providing specific information and does not imply recommendation or endorsement by the U.S. Department of Agriculture. USDA is an equal opportunity provider and employer.
- 1.Centers for disease control and prevention - CDC (2012) Pathogens causing US foodborne illnesses, hospitalizations, and deaths, 2000–2008 https://www.cdc.gov/foodborneburden/PDFs/pathogens-complete-list-01-12.pdf. Accessed 10 Apr 2017
- 2.Andrews WH, Wang H, Jacobson A, Hammack TS (2016) Salmonella. In: FOOD AND DRUG ADMINISTRATION. Bacteriological analytical manual (BAM) on line. Chap. 5. https://www.fda.gov/Food/FoodScienceResearch/LaboratoryMethods/ucm070149.htm>. Accessed 12 Apr 2017
- 9.Skladal P, Kovar D, Krajicek V, Siskova P, Pribyl J, Svabenska E (2013) Electrochemical immunosensors for detection of microorganisms. Int J Electrochem Sci 8(2):1635–1649Google Scholar
- 17.Luczak T, Osinska M (2017) New self-assembled layers composed with gold nanoparticles, cysteamine and dihydrolipoic acid deposited on bare gold template for highly sensitive and selective simultaneous sensing of dopamine in the presence of interfering ascorbic and uric acids. J Solid State Electrochem 21(3):747–758CrossRefGoogle Scholar
- 23.Green AA, Hughs WL (1955) Methods in enzymology, vol v. 1. Academic Press, New YorkGoogle Scholar
- 47.Knirel YA, Kocharova NA, Bystrova OV, Katzenellenbogen E, Gamian A (2002) Structures and serology of the O-specific polysaccharides of bacteria of the genus Citrobacter. Arch Immunol Ther Exp 50(6):379–391Google Scholar
- 48.Péterfi Z, Kustos I, Kilár F, Kocsis B (2007) Microfluidic chip analysis of outer membrane proteins responsible for serological cross-reaction between three gram-negative bacteria: Proteus Morganii O34, Escherichia Coli O111 and Salmonella Adelaide O35. J Chromatogr A 1155(1):214–217CrossRefGoogle Scholar