OVA is the predominant protein found in egg white, known for its functional properties, particularly its ability to foam and form gels. It is a globular protein with a molecular weight of 43 kDa, consisting of 385 amino acid residues, 1 disulfide bond, and 4 free sulfhydryl groups. Being a globular protein, OVA can undergo modifications through various treatments, including heating, high pressure, and enzymatic reactions.
OVA can be used as a protein carrier for polysaccharide vaccines. It is widely used as a protein carrier in tumor-associated carbohydrate antigens (TACAs).
Fig.1 Development of Eucommia ulmoides leaf polysaccharide-OVA vaccine. (Feng, et al., 2021)
CD BioGlyco provides OVA conjugation services with various polysaccharides, including capsular polysaccharides, polysaccharides extracted from organisms, and synthetic polysaccharides. Our polysaccharide-OVA conjugation service can refer to the following steps.
Polysaccharides are obtained by extraction, synthesis, or commercial means.
At first, the polysaccharide is activated. The polysaccharide is dissolved in sodium chloride solution and reacted with cyanogen bromide. Then, ADH solution is added and the pH of the reaction mixture is adjusted using a hydrochloric acid solution. The next day, the solution is dialyzed against EMS buffer to remove excess ADH and cyanogen bromide. Then the activated polysaccharide solution is mixed with OVA, and N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC) is added for reaction. Excess EDC is removed using a dialysis bag in NaCl solution to obtain the conjugated product of polysaccharide and OVA.
The separation and purification of the conjugate of polysaccharide and OVA is performed by column chromatography, the isolated and purified conjugate is freeze-dried, and the resulting powder is stored.
Polysaccharide-OVA conjugates are identified using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in stacking and resolving gels.
Polysaccharide-OVA conjugates are separated and purified using column chromatography. The BCA method is used to draw a standard curve with OVA as the protein standard to determine the protein content of the samples. After separation and purification, the quality of the polysaccharide-OVA conjugate is determined using a standard curve. The mass of polysaccharides in the polysaccharide-protein conjugate is determined by the phenol-sulfate method.
After adjusting the cell concentration of macrophages, place them in a 96-well cell culture plate. After culturing for 24 h, polysaccharide-protein conjugate solutions of different concentrations are added and cultured. Next, the reagent is added to each well and OD is measured later using a microplate reader.
Fig.2 Steps of polysaccharide-OVA conjugation development. (CD BioGlyco)
CD BioGlyco is committed to high-quality Carbohydrate-based Vaccine Development, and we provide a variety of solutions to assist your research. According to your needs, we provide you with a comprehensive one-stop OVA-polysaccharide conjugation service. If you are interested in our services, please contact us for more details without any hesitation.
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