2 atcc htb 37

Two L. monocytogenes strains (LM2 and LM9) with different ability to produce biofilm were selected. Both strains were obtained from a collection of strains of Microbiology Institute, Department of Public Health and Infectious Diseases, “Sapienza” University of Rome.
Strains were biochemically controlled by the API Listeria kit (Bio Mérieux, France), according to the manufacturer's instructions. Haemolysis on Muller Hinton agar (Oxoid) supplemented with 5% sheep blood was used as additional test. Bacteria were maintained as stock cultures in 15% glycerol-brain heart infusion broth (BHI) (Oxoid) at −80°C.
LM2 and LM9 strains were previously characterised for biofilm formation and classified as moderate and strong biofilm producers, respectively [28] .
Cells derived from a human colon carcinoma (CaCo-2) (ATCC® HTB-37) were purchased from American Type Culture Collection (ATCC; Manassas, VA, USA) and cultured in Dulbecco-modified minimum essential medium with Earle's salts (D-MEM, EuroClone), supplemented with 10% (v/v) heat-inactivated foetal calf serum (FCS, JRH Biosciences), and 2 mM glutamine. All incubations were carried out in a 5% CO₂ atmosphere at 37°C. Cells were used 48 hrs after seeding.

Human colon epithelial cells, Caco-2 (ATCC^®HTB-37™), were purchased from the American Type Culture Collection (Rockville, MD, USA). In our experiments, the Caco-2 cells were cultured in Dulbecco’s Minimal Essential Medium (DMEM) supplemented with 10 mM nonessential amino acids, 10% (v/v) fetal bovine serum (FBS), 100 μg/mL streptomycin, 100 U/mL penicillin, and 2 mM glutamine (growth medium). Cells were incubated at 37 °C and 5% CO₂, and sub-cultured at 80–90% confluence every 3–4 days.
For differentiation, Caco-2 cells were seeded at a density of 1 × 10⁵ cells/well in 12-well trans-well plate (12 mm, with 0.4 µm pore polycarbonate membrane Insert, Corning) and differentiated for 21 days in DMEM growth medium. The medium was replaced every 2–3 days for both the apical (AP) and basal (BL) sides of the trans-well filters. The integrity of cell monolayer was checked by measuring the trans-epithelial electrical resistance (TEER) before and after the experiments with an Epithelial Volt/Ohm Meter (EVOM).