title: Development of a Polyclonal Antibody-Based Sandwich Enzyme-Linked Immunosorbent Assay for the Detection of Spores of Alicyclobacillus acidoterrestris in Various Fruit Juices.
abstract: A polyclonal rabbit antibody-based sandwich ELISA for the rapid and specific detection of spores of Alicyclobacillus acidoterrestris was established. The reactivity of the antisera with spores was confirmed by immunofluorescence. For a thorough evaluation of the ELISA, 61 strains and isolates of Alicyclobacillus spp. were characterized regarding their guaiacol production ability and genetic variability. The ELISA was highly sensitive, the detection limits were isolate-dependent and ranged from 2.1 × 10(3) - 3.8 × 10(4) spores/mL, except for one isolate, for which a slightly lower sensitivity (5 × 10(5) spores/mL) was observed. Inclusivity tests revealed that the ELISA reacts with all tested A. acidoterrestris, while no cross-reactions with spores of 30 strains of Bacillus spp. and Clostridium spp. were observed. Further on, the assay applicability was tested with orange, apple (clear and unfiltered), tomato, pink grapefruit, pear, and white grape juices. Juices were inoculated with 1 or 10 spores/mL of A. acidoterrestris. After enrichment for 48 h, the established ELISA enabled the reliable and reproducible detection of contaminated samples. The enriched samples could be applied directly to the assay, underlining the robustness of the developed ELISA method.
authors.affiliation: Department of Veterinary Sciences, Faculty of Veterinary Medicine, Ludwig-Maximilians-Universität München , Schönleutnerstr. 8, 85764 Oberschleißheim, Germany.
authors.forename: Sophia
authors.initials: S
authors.lastname: Mast
authors.affiliation: Department of Veterinary Sciences, Faculty of Veterinary Medicine, Ludwig-Maximilians-Universität München , Schönleutnerstr. 8, 85764 Oberschleißheim, Germany.
authors.forename: Richard
authors.initials: R
authors.lastname: Dietrich
authors.affiliation: Department of Veterinary Sciences, Faculty of Veterinary Medicine, Ludwig-Maximilians-Universität München , Schönleutnerstr. 8, 85764 Oberschleißheim, Germany.
authors.forename: Andrea
authors.initials: A
authors.lastname: Didier
authors.affiliation: Department of Veterinary Sciences, Faculty of Veterinary Medicine, Ludwig-Maximilians-Universität München , Schönleutnerstr. 8, 85764 Oberschleißheim, Germany.
authors.forename: Erwin
authors.initials: E
authors.lastname: Märtlbauer
mesh_terms: D056450:Alicyclobacillus / Q000737:chemistry / Q000254:growth & development*; D000818:Animals; D015169:Colony Count, Microbial; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005506:Food Contamination / Q000032:analysis*; D000067030:Fruit and Vegetable Juices / Q000032:analysis*; D011817:Rabbits; D013171:Spores, Bacterial / Q000737:chemistry* / Q000254:growth & development
keywords: Alicyclobacillus acidoterrestris; detection; fruit juice; immunofluorescence; polyclonal antibody; sandwich ELISA; spores