Fluorescent Nanoparticle Interactions with Streptococcus pyogenes

S. pyogenes at ~10⁹ Colony Forming Unit (CFU)/mL, was incubated without or with 10% v/v of OFNDs, OLNDs, Ery-OFNDs, Ery-OLNDs—previously labeled with 7% of fluorescein isothiocyanate (FITC, Sigma-Aldrich)—for different incubation times and in agitation at 37 °C [24 (link),32 (link)]. After 3 or 24 h of incubation, bacteria were harvested by centrifugation at 3000× g at RT for 10 min, washed 2–3× with phosphate-buffered saline (PBS 1x) to remove unbound NDs. After the final wash, each sample was resuspended in 1 mL of PBS 1x and 50 μL of all the NDs were transferred onto separate glass slides, heat-fixed, and then stained for 15 min with 5 μg/mL of a propidium iodide (Invitrogen-Thermo Fisher Scientific Inc., Waltham, MA, USA) solution in a humid chamber at 37 °C. S. pyogenes was fixed with mounting solution and covered by cover slips. All ND formulations were observed using an Olympus IX70 inverted laser scanning confocal microscope, and images were recorded by using a FluoView 300 software (Olympus Biosystems, Melville, NY, USA) [24 (link),32 (link)].