The use of regenerable, affinity ligand-based surfaces for immunosensor applications.
The regeneration of antibody-binding surfaces is of major importance for re-usable sensor formats such as required for direct 'real-time' biosensing technologies and is often difficult to achieve. Antibodies commonly bind the antigen with high avidity and may themselves be sensitive to regeneration conditions. The interaction of polyclonal anti-chlorpyriphos antibody with an immobilised chlorpyriphos-ovalbumin (chlor-oval) conjugate and the interaction of soluble recombinant CD4 with covalently immobilised anti-CD4 IgG are presented in order to highlight these difficulties. Affinity-capture is suggested as an alternative format as it facilitates surface regeneration, directed immobilisation and the attainment of interaction progress curves that conform to the ideal pseudo-first-order kinetic interaction model. Protein A, protein G and polyclonal anti-mouse Fe-coated surfaces were used to observe the interaction of captured anti-GST monoclonal antibody with glutathione-s-transferase (GST). It was shown that a protein A affinity-capture surface produced ideal interaction progress curves while both protein G and polyclonal anti-mouse Fe resulted in systemic deviations.
Quinn
J
J
School of Biotechnology, Dublin City University, Glasnevin, Ireland.
Patel
P
P
Fitzpatrick
B
B
Manning
B
B
Dillon
P
P
Daly
S
S
Okennedy
R
R
Alcocer
M
M
Lee
H
H
Morgan
M
M
Lang
K
K
D000345:Affinity Labels; D000818:Animals; D000937:Antigen-Antibody Reactions; D015374:Biosensing Techniques / Q000379:methods*; D015704:CD4 Antigens / Q000276:immunology; D004390:Chlorpyrifos / Q000276:immunology; D005982:Glutathione Transferase / Q000276:immunology; D066298:In Vitro Techniques; D007700:Kinetics; D008024:Ligands; D051379:Mice; D009419:Nerve Tissue Proteins; D013205:Staphylococcal Protein A
D000345:Affinity Labels; D015704:CD4 Antigens; C029152:G-substrate; D008024:Ligands; D009419:Nerve Tissue Proteins; D013205:Staphylococcal Protein A; D005982:Glutathione Transferase; D004390:Chlorpyrifos