Liposome-mediated enhancement of the sensitivity in immunoassays of proteins and peptides in surface plasmon resonance spectrometry.
A recently developed liposome sandwich immunoassay for interferon-gamma (IFN-gamma), to be applied in microtiter plates, is tailored for surface plasmon resonance (SPR) spectrometry. The assay is performed on a thin (approximately 20 nm) polystyrene layer that covers a gold surface. This way, analytical data obtained from microtiter plate technology can directly be extrapolated toward SPR. For assaying the antigen IFN-gamma, a 16-kDa cytokine, a capture monoclonal antibody is physically adsorbed onto the polystyrene surface. After addition of the sample containing IFN-gamma, a biotinylated detecting antibody is added. Avidin is used as a bridging molecule between the biotinylated antibody and the biotinylated liposomes. All solutions are prepared with PBS buffer (10 mM, pH 7.4). This avoids additional changes in index of refraction caused by the use of various buffer solutions in immunoassays on microtiter plates for coating, binding, and washing procedures. It is shown that, when liposomes are used, a substantial enhancement of the detection limit is achieved. The "liposome" strategy improves the sensitivity for the IFN-gamma assay approximately 4 x 10(4) times and the detection limit to low picomolar. The method is generally applicable to other sandwich immunoassays.
Wink
T
T
Department of Pharmaceutical Analysis, Faculty of Pharmacy, Utrecht University, The Netherlands.
van Zuilen
S J
SJ
Bult
A
A
van Bennekom
W P
WP
D000327:Adsorption; D000911:Antibodies, Monoclonal / Q000276:immunology; D001360:Avidin; D019921:Biotinylation; D002138:Calibration; D007118:Immunoassay / Q000379:methods; D007371:Interferon-gamma / Q000032:analysis* / Q000276:immunology; D007834:Lasers; D008081:Liposomes; D010455:Peptides / Q000032:analysis*; D011137:Polystyrenes; D012680:Sensitivity and Specificity; D013057:Spectrum Analysis / Q000295:instrumentation / Q000379:methods
D000911:Antibodies, Monoclonal; D008081:Liposomes; D010455:Peptides; D011137:Polystyrenes; D001360:Avidin; D007371:Interferon-gamma