TY - JOUR
T1 - A comparative study on antibody immobilization strategies onto solid surface
AU - Lee, Ji Eun
AU - Seo, Jeong Hyun
AU - Kim, Chang Sup
AU - Kwon, Yunkyeoung
AU - Ha, Jeong Hyub
AU - Choi, Suk Soon
AU - Cha, Hyung Joon
PY - 2013/10
Y1 - 2013/10
N2 - Antibody immobilization onto solid surface has been studied extensively for a number of applications including immunoassays, biosensors, and affinity chromatography. For most applications, a critical consideration regarding immobilization of antibody is orientation of its antigen-binding site with respect to the surface. We compared two oriented antibody immobilization strategies which utilize thiolated-protein A/G and thiolated-secondary antibody as linker molecules with the case of direct surface immobilization of thiol-conjugated target antibody. Antibody immobilization degree and surface topography were evaluated by surface plasmon resonance and atomic force microscope, respectively. Protein A/G-mediated immobilization strategy showed the best result and secondary antibody-mediated immobilization was the worst for the total immobilization levels of target antibodies. However, when considering real-to-ideal ratio for antigen binding, total target antigen binding levels (oriented target antibody immobilization levels) had the following order: secondary antibody-mediated immobilization>protein A/G-mediated immobilization>direct thiol-conjugated immobilization. Thus, we confirmed that protein A/G- and secondary antibody-mediated strategies, which consider orientation of target antibody immobilization, showed significantly high antigen binding efficiencies compared to direct random immobilization method. Collectively, the oriented antibody immobilization methods using linker materials could be useful in diverse antibody-antigen interaction-involved application fields.
AB - Antibody immobilization onto solid surface has been studied extensively for a number of applications including immunoassays, biosensors, and affinity chromatography. For most applications, a critical consideration regarding immobilization of antibody is orientation of its antigen-binding site with respect to the surface. We compared two oriented antibody immobilization strategies which utilize thiolated-protein A/G and thiolated-secondary antibody as linker molecules with the case of direct surface immobilization of thiol-conjugated target antibody. Antibody immobilization degree and surface topography were evaluated by surface plasmon resonance and atomic force microscope, respectively. Protein A/G-mediated immobilization strategy showed the best result and secondary antibody-mediated immobilization was the worst for the total immobilization levels of target antibodies. However, when considering real-to-ideal ratio for antigen binding, total target antigen binding levels (oriented target antibody immobilization levels) had the following order: secondary antibody-mediated immobilization>protein A/G-mediated immobilization>direct thiol-conjugated immobilization. Thus, we confirmed that protein A/G- and secondary antibody-mediated strategies, which consider orientation of target antibody immobilization, showed significantly high antigen binding efficiencies compared to direct random immobilization method. Collectively, the oriented antibody immobilization methods using linker materials could be useful in diverse antibody-antigen interaction-involved application fields.
KW - Antibody Immobilization
KW - Atomic Force Microscopy
KW - Orientation
KW - Protein A/G
KW - Secondary Antibody
KW - Surface Plasmon Resonance
KW - Thiolation
UR - http://www.scopus.com/inward/record.url?scp=84884986326&partnerID=8YFLogxK
U2 - 10.1007/s11814-013-0117-5
DO - 10.1007/s11814-013-0117-5
M3 - Article
AN - SCOPUS:84884986326
SN - 0256-1115
VL - 30
SP - 1934
EP - 1938
JO - Korean Journal of Chemical Engineering
JF - Korean Journal of Chemical Engineering
IS - 10
ER -