Journal: Vaccine

Article Title: Saccharomyces cerevisiae as a platform for vaccination against bovine mastitis.

doi: 10.1016/j.vaccine.2024.126385

Figure Lengend Snippet: Fig. 2. – Production of a model yeast-based vaccine. A. Diagram depicting pYD1 and pYD1-OVA constructs. B. Representation of Aga2p-OVA expression by yeast surface display. C. Flow-cytometry analysis of OVA surface expression by EBY100-OVA after galactose (GAL) induction and heat-inactivation. EBY100-OVA non- inactivated or grown in raffinose (RAF) were used as controls. D. Western-blot analysis of OVA expression by EBY100-OVA. EBY100 was used as a negative con- trol. E. Analysis of OVA surface expression by EBY100-OVA after GAL induction and heat-inactivation by immunofluorescence using a rabbit anti-OVA polyclonal antibody. OVA staining is highlighted in red. Red bars correspond to 5 μm. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: For immunofluorescence, EBY100-OVA cultivated in the presence of galactose or raffinose (heat-inactivated at 56 ◦C for 30 min) was incubated in suspension with a rabbit polyclonal antibody anti-OVA antibody (produced by our team, 5 μg/ml) and a donkey anti-rabbit IgG (H + L) secondary antibody conjugated to Alexa Fluor 594 (Jackson ImmunoResearch, 1:100) diluted in FACS buffer.

Techniques: Construct, Expressing, Flow Cytometry, Western Blot, Immunofluorescence, Staining

Journal: Vaccine

Article Title: Saccharomyces cerevisiae as a platform for vaccination against bovine mastitis.

doi: 10.1016/j.vaccine.2024.126385

Figure Lengend Snippet: Fig. 3 – In. vivo evaluation of a yeast-based vaccine against mastitis. A. Scheme of immunisation with EBY100-OVA. B. Monitoring of rectal temperature after prime. C.. Rectal temperature and mammary gland evaluation after booster. D-E. PBMC from vaccinated animals were isolated in the indicated days and stimulated in vitro with empty EBY100 or OVA. IFNγ (D) and IL-17 (E) release in supernatants was measured by ELISA. F. Analysis of serum total antibodies anti-EBY100 and anti- OVA by ELISA. G. Scheme depicting intramammary stimulations of cows immunised as described in A with recombinant OVA. H. Somatic cells count in mammary gland secretion before and after intramammary stimulations. I. Scheme of immunisation with adjuvanted-recombinant OVA. J-K. PBMC from OVA (J) or EBY100- OVA (K) immunised cows were stimulated in vitro with empty EBY100, EBY100-OVA or OVA. IFNγ and IL-17 release was analysed by ELISA. Data shown in B–F, H and K were obtained from the same cows immunised with EBY100-OVA (A) and subsequently stimulated via the intramammary route with recombinant OVA (G). Asterisks denote statistically significant difference (indicated time points versus day 0).

Article Snippet: For immunofluorescence, EBY100-OVA cultivated in the presence of galactose or raffinose (heat-inactivated at 56 ◦C for 30 min) was incubated in suspension with a rabbit polyclonal antibody anti-OVA antibody (produced by our team, 5 μg/ml) and a donkey anti-rabbit IgG (H + L) secondary antibody conjugated to Alexa Fluor 594 (Jackson ImmunoResearch, 1:100) diluted in FACS buffer.

Techniques: In Vivo, Isolation, In Vitro, Enzyme-linked Immunosorbent Assay, Recombinant