polyclonal rabbit anti ova antibody  (Bio-Rad)

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

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Highly Sensitive, Flow Cytometry-Based Measurement of Intestinal Permeability in Models of Experimental Colitis

doi: 10.1016/j.jcmgh.2022.10.004

Figure Lengend Snippet: Muc2 and its glycosylation are critical for limiting OVA interactions with the intestinal epithelium and regulating intestinal permeability. ( A ) Visualization of OVA localization in the colon. Colonic tissue samples containing fecal matter from C57BL/6 mice were harvested and fixed with methyl-Carnoy’s fixative, and then stained for OVA (red), fucosylated residues on mucins (Ulex Europaeus Agglutinin I [UEA-1] lectin, green), epithelial cells (E-cadherin, white), and nuclei (DAPI, blue). ( B ) OVA or FITC-D assay readouts form the plasma samples of Muc2 +/+ (WT) (n = 6), Muc2 +/- (n = 7), and Muc2 -/- mice (n = 6) co-administered OVA and FITC-D. Data are representative of at least 3 independent experiments. Statistical significance was determined by 1-way analysis of variance, using the Tukey post hoc test. ( C ) A cohort of 10-week-old female Muc2 -/- mice (n = 5) was gavaged with 1 mg/mouse OVA as indicated and 2.5 μL blood samples were taken as indicated to track intestinal permeability changes within each animal. ( D ) Mechanistic actions of core 1 and core 3 synthases in the glycosylation of mucins. ( E ) Representative intestinal permeability data using the OVA assay on 8- to 10-week-old IEC– C1galt1 -/- mice (n = 5) and ( F ) C3GnT -/- mice (n = 3) (denoted Core 1 -/- and Core 3 -/- mice, respectively) relative to control IEC– C1galt1 fl/fl ( Core 1 fl/fl ) (n = 5) and C57BL/6 mice (n = 5). ( G ) Comparison of intestinal permeability from OVA assay readouts between IEC– C1galt1 -/- and C3GnT -/- mice in panels E and F 1 hour after OVA gavage. Mice were gavaged with 1 mg/mouse OVA and 2.5 μL blood samples were taken at the indicated time points. Data are representative of 3 separate experiments. C1GALT1, core 1 β1,3-galactosyltransferase; DAPI, 4′,6-diamidino-2-phenylindole; Gal, galactose; GalNAc, N-acetylgalactosamine; GlcNAc, N-acetylglucosamine; S, serine; T, threonine; β3Gn-T6, β1,3-N-acetylglucosaminyltransferase 6. ∗ P ≤ .05, ∗∗ P ≤ .01, ∗∗∗ P ≤ .001.

Article Snippet: For visualizing of OVA crossing the IEC barrier, rabbit anti-OVA polyclonal antibody (#GTX21221, 1:1000; GeneTex) or goat anti-OVA polyclonal antibody (#0855303, 1:100; MP Biomedicals) and mouse monoclonal anti-mouse E-cadherin antibody (#610182, 1:400; BD Transduction Laboratories) were used.

Techniques: Glycoproteomics, Permeability, Staining, Clinical Proteomics, Control, Comparison