A practical kinetic model for efficient isolation of useful antibodies from phage display libraries

• Published in: Journal of molecular catalysis. B, Enzymatic Enzymatic, 2004-06, Vol.28 (4), p.191-200
• Main Authors: Katakura, Yoshio, Zhuang, Guoqiang, Nakatani, Tomohiro, Furuta, Tetsuo, Omasa, Takeshi, Kishimoto, Michimasa, Suga, Ken-ichi, Shioya, Suteaki
• Format: Article
• Language: English
• Subjects: Antibody; Association rate constant; Kinetic model; On-rate selection; Phage display
• ISSN: 1381-1177; 1873-3158
• DOI: 10.1016/j.molcatb.2004.02.016

To isolate phages displaying a practical and useful antibody with a high k on value and/or a low k off value from phage display antibody libraries, we developed a rational strategy based on a kinetic model. In the model, the recovery of a phage displaying an antibody after a round of biopanning is expressed as a function of five parameters, the apparent association rate constant of the phage antibody to the immobilized antigen ( k on′), the apparent dissociation rate constant of the phage antibody from the immobilized antigen ( k off′), the effective antigen concentration ( C), the time for the binding process ( t b) and the time for the washing process ( t w). An optimum set of operating parameters ( C, t b and t w) for isolating phages displaying an antibody with a high k on value was designed based on the model. Three rounds of biopanning were carried out under the designed conditions, against a phage library in which the hypervariable regions of an original antibody were randomized. All isolated phages displayed an antibody with a higher k on value and one displayed an antibody with a 30-fold greater k on value than that of the original antibody. Experimental conditions which improve the efficiency of conventional off-rate selections are also described.