Sigma 500 vp field emission scanning electron microscope

A small droplet (1–2 μL) of either SLN or SLN-Nisin were allowed to dry on glass coverslips overnight under vacuum. The next day, the dried samples were sputter coated with a 1 nm fine grained Pd–Au layer on an EM ACE600 sputter coater (Leica, Germany) and evaluated under a Sigma 500-VP Field-Emission Scanning Electron Microscope (SEM) (Zeiss, Germany). The average diameter was calculated using ImageJ and based on the images obtained. PDI was obtained by the following equation, according to Raval et al. [35 ]:
$$\text{PDI}={\left(\sigma /d\right)}^2$$
Where d represents the average diameter of the nanoparticles and σ represents their standard deviation.

HSC-3 cells were seeded in 12-well plates (1 × 10⁶ cells/well) containing glass cover slips and allowed to adhere overnight. HSC-3 cells were challenged with 100 μg/mL of Nisin, SLN-Nisin or an equivalent volume of SLN for 24 h.) were added to the cells for 24 h. After this, cells were rinsed with PBS and 10% glutaraldehyde for 20 min at room temperature and dehydrated using increasing ethanol solutions (30%, 50%, 70%, 85% and 95% v/v) for 10 min each. The next day, samples were sputter coated with a 1 nm fine-grained Pd–Au layer on an EM ACE600 sputter coater (Leica, Germany) and evaluated under a Sigma 500-VP Field-Emission Scanning Electron Microscope (SEM) (Zeiss, Germany).