title: Liposome-based immunostrip for the rapid detection of Salmonella.
abstract: Salmonellae are ubiquitous human pathogens, which pose a danger to the elderly and children. Due to the increased number of outbreaks of human illness associated with the consumption of contaminated products in the USA and many other countries, there is an urgent need to develop rapid assays to detect common food-borne pathogens. This study demonstrates the feasibility of using a detectable label comprising methyl blue (MB), a visible dye, entrapped inside liposomes. Immunoliposomes tagged with anti-Salmonella common structural antigens (CSA) antibody encapsulating MB dye were prepared and used as the signal amplifier for the development of a field-portable colorimetric immunoassay to detect Salmonellae. Tapping mode atomic force microscopy (TMAFM), a scanning probe technique, was utilized to demonstrate the presence of anti-Salmonella antibody at the thus-prepared liposome. A plastic-backed nitrocellulose strip with two immobilized zones formed the basis of a sandwich assay. The first zone was the antigen capture zone (AC zone), used in a sandwich (noncompetitive) assay format; the other was the biotin capture zone (BC zone), used as a quality control index for the strip assay. During the capillary migration of the wicking reagent containing 80 μL of immunoliposomes and 40 μL of the test sample (heat-killed S. typhimurium), sample pathogens with surface-bound immunoliposomes were captured at the AC zone, while the unbound immunoliposomes continued to migrate and bind to the anti-biotin antibodies coated on the BC zone. The color density of the AC zone was directly proportional to the number of Salmonella typhimurium in the test sample. The detection limit of the current assay with heat-killed Salmonella typhimurium was 1,680 cells. The cross-reactivity of the proposed immunoassay was also investigated, and pathogens including E. coli O157:H7 and Listeria genus specific caused no interference with the detection of Salmonella typhimurium.
authors.affiliation: Department of Chemistry, National Tsing Hua University, 101, Section 2, Kuang-Fu Road, Hsinchu, 30013, Taiwan. jaho@mx.nthu.edu.tw
authors.forename: Ja-An Annie
authors.initials: JA
authors.lastname: Ho
authors.forename: Shi-Chin
authors.initials: SC
authors.lastname: Zeng
authors.forename: Wei-Hsiang
authors.initials: WH
authors.lastname: Tseng
authors.forename: Yong-Jen
authors.initials: YJ
authors.lastname: Lin
authors.forename: Chun-Hsien
authors.initials: CH
authors.lastname: Chen
mesh_terms: D000368:Aged; D000907:Antibodies, Bacterial / Q000032:analysis; D000942:Antigens, Bacterial / Q000737:chemistry; D001557:Benzenesulfonates / Q000737:chemistry; D001710:Biotin / Q000737:chemistry; D002648:Child; D003101:Collodion / Q000737:chemistry; D003429:Cross Reactions; D019453:Escherichia coli O157 / Q000276:immunology; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D008081:Liposomes / Q000648:ultrastructure*; D008087:Listeria / Q000276:immunology; D018625:Microscopy, Atomic Force; D012480:Salmonella Infections / Q000175:diagnosis / Q000382:microbiology; D012486:Salmonella typhimurium / Q000276:immunology* / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D013194:Staining and Labeling
chemical_list: D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001557:Benzenesulfonates; D008081:Liposomes; D001710:Biotin; D003101:Collodion; C414357:Methyl Blue