Fig. 3: Glycan chain length depends on a number of factors and conditions applied during glycoconjugate vaccine production.

a Standard glycoconjugate vaccine production process. b Bioconjugation process. Critical junctures are indicated by numbers. 1. Genetics of the bacterial strain employed. The bioconjugation process includes all genetic modifications introduced to the glycoconjugate production strain to facilitate in vivo coupling of the glycan to a carrier protein. 2. Growth conditions. 3. Strategies for isolation and purification of the glycan (or, in bioconjugation, of the conjugate). 4. Size-exclusion-based glycan or glycoconjugate purification procedures, such as tangential filtration (TFF) or size-exclusion chromatography. 5. Chemical or physical glycan size-reduction protocols, such as acid hydrolysis, ozonolysis, microfluidization, and/or periodate oxidation. 6. Activation chemistries of glycan and carrier protein. 7. Conditions applied for the chemical coupling of carrier and glycan. 8. Purification via combinations of chromatographic techniques, such as ion exchange and hydrophobic interaction.