To cause infections, bacteria must colonize their host. Bacterial pathogens express various molecules or structures able to promote attachment to host cells 1 . These adhesins rely on interactions with host cell surface receptors or soluble proteins acting as a bridge between bacteria and host. Adhesion is a critical first step prior to invasion and/or secretion of toxins, thus it is a key event to be studied in bacterial pathogenesis. Furthermore, adhered bacteria often induce exquisitely fine-tuned cellular responses, the studies of which have given birth to the field of 'cellular microbiology' 2 . Robust assays for bacterial adhesion on host cells and their invasion therefore play key roles in bacterial pathogenesis studies and have long been used in many pioneer laboratories 3,4 . These assays are now practiced by most laboratories working on bacterial pathogenesis.Here, we describe a standard adherence assay illustrating the contribution of a specific adhesin. We use the Escherichia coli strain 2787 5 , a human pathogenic strain expressing the autotransporter Adhesin Involved in Diffuse Adherence (AIDA). As a control, we use a mutant strain lacking the aidA gene, 2787ΔaidA (F. Berthiaume and M. Mourez, unpublished), and a commercial laboratory strain of E. coli, C600 (New England Biolabs). The bacteria are left to adhere to the cells from the commonly used HEp-2 human epithelial cell line. This assay has been less extensively described before 6 . Protocol Preliminary: Bacterial strains and epithelial cells.Manipulations of cells and bacteria are performed aseptically, under a laminar flow hood. , and C600 from glycerol stocks on Lysogeny Broth (LB) agar plates (1% tryptone, 0.5% sodium chloride, 0.5% yeast extract, 1.5% agar) and grow at 37°C. To minimize variability in the assay, it is advised to always use freshly plated strains and to keep the strains at 4°C on sealed Petri plates only for a maximum of a couple of weeks. 2. Culture HEp-2 cells (ATCC CCL-23) in high glucose Dulbecco's Modified Eagle Medium (DMEM) supplemented with 10% heat inactivated bovine serum (heat inactivation is performed at 56°C for 30 min). During routine culture we also add 10 U/ml penicillin and 10 μg/ml streptomycin. 3. Grow Hep-2 cells at 37°C in a cell incubator with an atmosphere containing 5% CO2. Use standard cell culture procedures to maintain the cells, which are grown in 75 cm 2 flasks and subcultured every time they reach confluence. We use our cultured cells until they reach 30 to 40 passages and then discard them. 4. Prepare an assay when the HEp-2 cells in a 75 cm 2 flask are nearly reaching confluence and therefore are ready for an adherence assay. DAY 1: Preparing the inoculum and the epithelial cells DAY 2: Infection of cells1. Wash the HEp-2 cells in the flask once with warm Dulbecco's phosphate buffered saline (DPBS: 8 g/L NaCl, 0.2 g/L KCl, 0.2 g/L KH2PO4, 0.21 g/L Na2HPO4:7H2O) 2. The cells are incubated with 0.05% trypsin -EDTA for 5 min before adding fresh warm complete medium. After fresh medium i...

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The purpose of the study was to evaluate the reliability and validity of choice reaction time as a measure of recovery from general anesthesia. An experimental group of 43 patients underwent surgery under general anesthesia; they were measured before anesthesia and also 90, 150 and 210 min. after the end of anesthesia. A control group of 38 underwent the same procedure. Choice reaction time was not a valid measure of recovery from general anesthesia since the test was not reliable.