Comparison of Random and Site-Directed Immobilization of Antibody for α-Fetoprotein Direct Immunoassay by Protein Chip

Abstract

For immunoassay, immobilization of antibody is the most important technique in bioengineering for maintaining its intrinsic activity of antigen binding. The sugar chain in the Fc region carried by antibody can be oxidized into aldehyde group by sodium m-periodate, which can couple with the amino-group on chemistry modified substrate surface through covalent linking. Thereby, in situ site-directed immobilization of the oxidized antibody for only one-step reaction can be performed. Here we propose a facile method that oxidized anti-human α-fetoprotein antibody (anti-AFP) was in situ site-directed immobilized on APTES treated silicon substrate utilizing microfluidic channel system for AFP immunoassay. The shape of the immobilized anti-AFP was observed by AFM and imaged by imaging ellipsometry of self-developed protein system. Results shown that the amount of antibody immobilization amount and capacity of antigen binding of random immobilization and site-directed immobilization were separately enhanced 16.0% and 7.0%. the suggested strategy has the advantages of simple, environment-friend and convenient-operation.