Annual Report - Center for Food Safety Engineering - Purdue ...

Lu, Bhunia and ChengRapid, quantitative, and reusable immunosensors forbacteria detection on a microfluidic platformInvestigators: Chang Lu (Department of Agricultural and Biological Engineering), Arun Bhunia (Department ofFood Science), Zhongyang Cheng (Materials Engineering, Auburn University)Project RationalePortable, rapid, and sensitive biosensors for food safetyapplications enable point-of-care contamination detectionand immediate interpretation of the results. In our researchproject, we proposed to develop an integrated biosensorsystem on a microfl uidic chip for detecting bacteria basedon immunoassays. The device will offer a sensitivity of 10 2to 10 3 bacteria cell detection and an assay time of fewer than20 minutes for a single test. Our system will yield quantitativedata for estimating the number of the target bacterium in a foodsample. The microfl uidic system will consist of individual devicesfor cell lysis, lysate purifi cation, and immunoassays. In principle,the tool will be effective for any bacterium or strain given theavailability of a suitable intracellular antigen-antibody pair. In thisproject, we will demonstrate the concept using an intracellularantigen, alcohol acetaldehyde dehydrogenase (Aad), and itsantibody MAb-H7 to detect Listeria monocytogenes. In order toconcentrate L. monocytogenes cells from food samples, we willfabricate magnetic nanobars with different sizes and geometriesand develop protocols for immobilizing antibodies specifi c to L.monocytogenes on the surface.A portable, reusable, and low-cost device would be usefulfor point-of-care analysis in the food manufacturing industry.Conducting bacteria detection tests within food manufacturinglaboratories would dramatically decrease the turnaroundtime for the results and avoid potential contamination andchanges in the bacteria during transit. Conventional analyticalmethods require bulky, expensive equipment that are oftencost-prohibitive for food manufacturing laboratories. With ourlab-on-a-chip approach, sophisticated functions of a biologicallaboratory can be miniaturized on a microchip, enabling anyminimally equipped laboratory with the ability to perform bacteriadetection tests. This technology can signifi cantly benefi t the foodindustry by enhancing the laboratory-testing capabilities of foodmanufacturers and food testing laboratories as well as fi eldtestingactivities of governmental agencies.Project Objectives• Fabricate magnetic nanobars with different sizes andgeometries and develop protocols for immobilizingantibodies specifi c to L. monocytogenes on the surface.The amount of bacterial cells bound to the surfacewill be characterized under different conditions.• Develop an electrophoresis-based immunoassaycoupled with laser-induced fl uorescence on a microfl uidicchip. We will use this tool to quantitatively detect L.monocytogenes based on cell lysate via the interactionbetween alcohol acetaldehyde dehydrogenase(Aad) and its monoclonal antibody (MAb-H7).• Demonstrate a prototype-integrated microfl uidicsystem which incorporates different steps suchas manipulation of magnetic nanobars, cell lysis,lysate purifi cation, and immunoassay.Project HighlightsWe integrated the concentration, lysis, and competitiveimmunoassay for detecting L. monocytogenes on a microfl uidicchip. A packed bed of microbeads, with the bead surface coatedwith monoclonal antibody (MAb-H7) that is specifi c to the antigenListeria adhesion protein (LAP), was formed in a microfl uidicchannel to physically trap bacterial cells. Electrical lysis wasthen used to release intracellular materials from the trappedcells. Fluorescein isothiocyanate (FITC)-labeled LAP was fl owedthrough the microbead bed to bind to unreacted antibody sitesand reveal whether LAP was present in the bacterial sample. Byintegrating all these functions onto one simple portable chip, wecan produce a biosensor system that is both highly effi cient andinexpensive.12Center for Food Safety Engineering“By integrating all these functions onto one simple portable chip, we canproduce a biosensor system that is both highly effi cient and inexpensive.”