Image analysis software

The heights of mica-adsorbed HRP were determined as the heights corresponding to the maxima of the respective distributions of the mica-adsorbed objects with heights ρ(h) as reported in [41 (link)]:
where N_h is the number of AFM-visualized mica-adsorbed objects of the height h, while N is the total number of mica-adsorbed objects visualized by AFM.
The experimentally obtained dependence (1) was then approximated using the Gaussian function: $$\rho \left(h\right)={{\displaystyle \sum }}^{}{\rho }_i\left(h\right)={\displaystyle \sum }_{i=1}^2\frac{A{e}^{\frac{-4 ln(2){(h-h_c)}^2}{w^2}}}{w\sqrt{\frac{\pi }{4 ln(2)}}}$$
where A, h_c, and w are the parameters varied upon approximation. The maxima of the ρ(h) distributions were calculated as the maxima of the approximation Function (2) for each distribution. The analysis of the approximation of the experimentally obtained ρ(h) distributions is based on a χ² criterion.
The initial processing of the AFM images obtained (second-order plane subtraction) and the data export to ASCII format were performed with an Image Analysis software (NT-MDT, Zelenograd, Russia) supplied with the atomic force microscopes. The number of objects, visualized in the AFM images obtained, was calculated with a specialized Recognite software (developed in Institute of Biomedical Chemistry in collaboration of Moscow Engineering Physical Institute MEPhI).

Topography, phase, and magnitude signal images of the micelles deposited onto a freshly cleaved mica surface were obtained by atomic force microscopy (AFM) using a scanning probe microscope, NTEGRA Prima (NT-MDT, Moscow, Russia), operated in a semi-contact mode with a 15–20 nm peak-to-peak amplitude of the “free air” probe oscillations. Silicon cantilevers NSG01 “Golden” series cantilevers for semi-contact mode (NT-MDT, Russia) were used. Image processing was performed using the Image Analysis software (NT-MDT, Russia).

Chit5 and Chit5-PEG nanoparticles were obtained using extrusion (200 or 400 nm membrane, Avanti Polar Lipids) after 1 h incubation of samples (0.01–1 mg/L) at 40 °C. The initiation of gelation was performed by placing a solution with an alkaline medium (pH 8–10), ovalbumin or alginic acid as the counterions in a syringe receiver. Particles’ hydrodynamic diameter sizes and ζ-potentials were measured using Zetasizer Nano S from Malvern (Malvern, UK) (4 mW He–Ne laser, 633 nm, scattering angle 173°) in 0.01 M PBS (pH 7.4). Dynamic light scattering data were analyzed using Zetasizer Software (v. 8.02). Topography, phase and magnitude signal images of the micelles deposited onto freshly cleaved surface of mica were obtained via atomic force microscopy (AFM) using a scanning probe microscope, NTEGRA Prima (NT-MDT, Russia), operated in a semi-contact mode with 15–20 nm peak-to-peak amplitude of the “free air” probe oscillations. Silicon cantilevers NSG01 “Golden” series were used for semi-contact mode (NT-MDT, Russia). Image processing was performed using the Image Analysis software (NT-MDT, Russia).