Kinetic analysis of recombinant antibody-antigen interactions: relation between structural domains and antigen binding.
The relation between domain structures of recombinant monoclonal antibody fragments and their reaction kinetics was studied for the first time using a novel biosensor based on surface plasmon resonance technology. The association and dissociation rate constants of Fab, Fv and single domain (VH fragment) anti-lysozyme antibodies were determined and compared to the intact monoclonal antibody. Fab and Fv fragments showed similar reaction kinetics and had affinity constants of 6 x 10(9) M-1 and 25 x 10(9) M-1, respectively. The single domain antibody had significantly different reaction kinetics compared to the fragments consisting of paired heavy and light chain domains. The VH domain had both a higher dissociation and a lower association rate constant, which resulted in an affinity constant approximately 250 times lower than the Fab fragment. This rapid evaluation of antibody reaction kinetics should prove to be an important selection parameter when comparing antibody fragments for their utility in therapeutic or other applications.
Borrebaeck
C A
CA
Department of Immunotechnology, Lund University, Sweden.
Malmborg
A C
AC
Furebring
C
C
Michaelsson
A
A
Ward
S
S
Danielsson
L
L
Ohlin
M
M
D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology*; D000937:Antigen-Antibody Reactions; D001666:Binding Sites, Antibody; D015374:Biosensing Techniques; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry / Q000276:immunology*; D007128:Immunoglobulin Fragments / Q000737:chemistry / Q000276:immunology*; D007135:Immunoglobulin Variable Region / Q000737:chemistry / Q000276:immunology*; D007700:Kinetics; D009113:Muramidase / Q000276:immunology*; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology
D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D007128:Immunoglobulin Fragments; D007135:Immunoglobulin Variable Region; D011994:Recombinant Proteins; C055966:immunoglobulin Fv; D009113:Muramidase