Numerous techniques have been developed to prepare immunoliposomes based on the nucleophilic reactivity of free amine groups of proteins or peptides. One of the most popular and commonly used methods is to covalently couple free carboxylic groups to primary amines through activation of the carboxyl groups with EDC (1-ethyl-3-[3-dimethylaminopropyl] carbodiimide). EDC, which is a so-called zero-length crosslinking agent, reacts with the carboxyl to form an amine reactive intermediate (O-acylisourea). The produced O-acylisourea can be easily displaced by nucleophilic attack from the primary amino groups in the reaction mixture. However, this intermediate is unstable and hydrolyzed in aqueous solutions. In order to prevent the intermediate hydrolysis, sulfo-NHS (N-hydroxysulfosuccinimide) is added to EDC to produce a significantly more stable and more soluble active intermediate (NHS ester).
Consequently, the immunoliposomes are prepared by a two-step coupling procedure: first, activating the free carboxyl group of the linker lipid incorporated in the liposomes with EDC and sulfo-NHS, and then covalently conjugating the antibodies to the lipids through displacement of sulfo-NHS groups by antibody amines, as depicted below. EDC/sulfo-NHS coupling reactions are highly selective and highly efficient, and the biological activity of the protein or peptide is preserved.
ImmunoFluor™-Donecanyl is a non-PEGylated product. For other amine reactive (PEGylated and non-PEGyalated products) and also ImmunoFluor™ products suitable for other types conjugation methods see here.
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采用抗-HBe抗体包被反应板，加入校准品及被测样本，同时加入定量HBeAg中和抗原，经过振荡孵育,洗板后再加入铕标记的抗-HBe，若标本中抗-HBe浓度高，HBeAg将被大量中和，使最后形成的抗-HBe-HBeAg-铕标记抗-HBe复合物减少。增强液（β－NTA）将标记在抗体上的Eu3+解离到溶液中，Eu3+和增强液中的有效成分形成高荧光强度的螯合物，荧光强度和样本中的抗-HBe浓度成反比。
