title: Comparative detection of bacterial adhesion to Caco-2 cells with ELISA, radioactivity and plate count methods.
abstract: Different methods are used to study bacterial adhesion to intestinal epithelial cells, which is an important step in pathogenic infection as well as in probiotic colonization of the intestinal tract. The aim of this study was to compare the ELISA-based method with more conventional plate count and radiolabeling methods for bacterial adhesion detection. An ELISA-based assay was optimized for the detection of Bifidobacterium longum and Escherichia coli O157:H7, which are low and highly adherent bacteria, respectively. In agreement with previous investigations, a percentage of adhesion below 1% was obtained for B. longum with ELISA. However, high nonspecific background and low positive signals were measured due to the use of polyclonal antibodies and the low adhesion capacity with this strain. In contrast, the ELISA-based method developed for E. coli adhesion detected a high adhesion percentage (15%). For this bacterium the three methods tested gave similar results for the highest bacterial concentrations (6.8 Log CFU added bacteria/well). However, differences among methods increased with the addition of decreased bacterial concentration due to different detection thresholds (5.9, 5.6 and 2.9 Log CFU adherent bacteria/well for radioactivity, ELISA and plate count methods, respectively). The ELISA-based method was shown to be a good predictor for bacterial adhesion compared to the radiolabeling method when good quality specific antibodies were used. This technique is convenient and allows handling of numerous samples.
authors.affiliation: Laboratory of Food Biotechnology, Institute of Food Science and Nutrition, Swiss Federal Institute of Technology, Schmelzbergstrasse 9, CH-8092 Zürich, Switzerland.
authors.forename: Gwenaëlle
authors.initials: G
authors.lastname: Le Blay
authors.forename: Ismaïl
authors.initials: I
authors.lastname: Fliss
authors.forename: Christophe
authors.initials: C
authors.lastname: Lacroix
mesh_terms: D001422:Bacterial Adhesion / Q000502:physiology*; D001644:Bifidobacterium / Q000502:physiology*; D018938:Caco-2 Cells; D015169:Colony Count, Microbial; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D004927:Escherichia coli Infections / Q000473:pathology; D019453:Escherichia coli O157 / Q000502:physiology*; D006801:Humans; D007413:Intestinal Mucosa / Q000382:microbiology*; D012588:Scintillation Counting