Synthesis And Characterization Of Conjugates Essay

1653 Words Sep 20th, 2016 7 Pages
Synthesis and characterisation of conjugates
Conjugation of CpG to antigen as a means for co-delivery has been shown to enhance cellular immune responses compared to delivery as a mixture. The ODN CpG however can be conjugated via its 3’ or 5’ end, and blocking of the 5’ end has been described to reduce its immunostimmulatory properties in splenocytes. To determine whether the end of CpG conjugated to OVA effects the induction of a cellular immune response by an immunotherapeutic vaccine, CpG modified with an amine group on either the 3’ end or the 5’ end were conjugated to the model tumour antigen OVA. OVA was first purified from aggregates by preparative size-exclusion chromatography (SEC) and then conjugated onto CpG using a bis-aryl-hydrazone linking strategy 30. As shown in Figure 1, the terminal amine group at either the 3’ or 5’ end of CpG was modified to an aromatic aldehyde by reacting the terminal amine group with the activated ester of the linker succinimidyl 4-formylbenzoate (4FB). The average molar substitution ratio (MSR) of the CpGs with the 4FB linker was 1.0 ± 0.2. Next the lysine residues on OVA were modified to aromatic hydrazine groups by reaction with the activated ester of the linker succinimidyl 6-hydrazinonicotinate acetone hydrazone (HyNic). The average MSR of OVA with the HyNic linker was 6.1 ± 1.6. The HyNic modified OVA was reacted with either the 3’ or 5’ 4FB modified CpG at a molar ratio of 4:1 to form a stable covalent bis-aryl hydrazone bond.…

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