Experimental strategy and Piscirickettsia salmonis proteoliposome light scattering characterization. A Fish were left in acclimatization for 14 days and then were vaccinated intraperitoneally in freshwater. Salinity was increased gradually. 300 DD after first vaccination fish received a second vaccination. 300 DD later, fish were challenged with Piscirickettsia salmonis. Blood samples to quantify specific IgM were taken pre-vaccination, after the first immunization and second immunization (red squares). The experimental strategy can be separated into 3 steps: acclimatization (yellow), immunization (blue), challenge (green). DD, degree-days. B Proteoliposome zeta potential. C Proteoliposome size with a lognormal distribution. D Zeta potential shows an anionic proteoliposome with an average superficial charge of -37.9 ± 6.3 mV (n = 10). E Polydispersity index was 0.1349 ± 0.03. F The average diameter of proteoliposome was 250.9 ± 13.2 nm (n = 10).