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Superparamagnetic Nanoparticles Based Immunomagnetic Separation—Multiplex Polymerase Chain Reaction Assay for Detection of Salmonella

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We report a method that combines Immunomagnetic separation and polymerase chain reaction (IMS-PCR) to detect Salmonella sp. The IMS-PCR assay exploits selective extraction of bacteria using antibody coated superparamagnetic particles and multiplex PCR amplification that enable detection of Salmonella sp. The anti-Salmonella polysera used was raised in female BalbC mice using heat inactivated Salmonella whole cell. The common organosilane aminopropyltriethoxy silane (APTES) under reflux conditions amine functionalizes Fe2O3 nanoparticles enabling covalent attachment of anti-Salmonella polysera. We examine the efficiency to capture and detect target bacteria in suspension using immunomagnetic particles in two different attachment modes namely, random assembly and oriented assembly of antibodies. The functionalized Fe2O3 nanoparticles conjugated with Salmonella antibody specifically capture Salmonella sp. from bacterial suspension. Retrieved genomic DNA of captured bacteria were amplified using set of four primers specifically targeting the invA, fliC, viaB, and prt genes of Salmonella Typhi and Salmonella Paratyphi A by multiplex PCR. We find that site-directed attachment and random attachment of antibody was sensitive to 107 and 106 cells/ml respectively. Thus we demonstrate IMS-PCR with improved surface chemistry and covalent attachment of antibody was sensitive to 106 and 107 cells/ml respectively. Thus we demonstrate IMS-PCR with improved surface chemistry and covalent attachment of antibody to be rapid, specific and cost effective detection method for Salmonella species in clinical isolates.
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Keywords: ANTIBODY CONJUGATION; BACTERIAL DETECTION; FUNCTIONALIZED NANOPARTICLES; IMMUNOMAGNETIC SEPARATION; MULTIPLEX PCR; SALMONELLA SP

Document Type: Research Article

Publication date: 01 January 2012

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  • Science of Advanced Materials (SAM) is an interdisciplinary peer-reviewed journal consolidating research activities in all aspects of advanced materials in the fields of science, engineering and medicine into a single and unique reference source. SAM provides the means for materials scientists, chemists, physicists, biologists, engineers, ceramicists, metallurgists, theoreticians and technocrats to publish original research articles as reviews with author's photo and short biography, full research articles and communications of important new scientific and technological findings, encompassing the fundamental and applied research in all latest aspects of advanced materials.
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