Improved functionality of antibody-colloidal gold conjugates with the aid of lipoamide-grafted N-[tris(hydroxymethyl)methyl]acrylamide polymers.
Colloidal gold has been used as a label in sandwich assays for human IgG, in which intercalating N-[tris(hydroxymethyl)methyl]acrylamide (pTHMMAA) polymers have been employed to stabilise the particles coated with antibody fragments. A direct absorbance reading of the particles could be obtained from sandwich assays on polystyrene, and a strongly amplified response was observed in similar assays based on Surface Plasmon Resonance (SPR): for h-IgG, detection limits below 100 pg/mL could be achieved. Three different polymer lengths and two different particles sizes were compared in sandwich assays performed on polystyrene and gold. The resulting binding curves fitted well to the Langmuir-Freundlich isotherm and the binding constants were in good agreement with the values found in earlier studies. The amplification afforded by the nanoparticles was strongly dependent on the antigen concentration, on the type of polymer and on the particle size. Compared to the direct response of the antigen, amplification factors larger than 100 could be achieved. The study proves that the polymers give stabilised particles, which can be used in highly sensitive sandwich assays.
Albers
Willem M
WM
VTT Technical Research Centre of Finland, Molecular Sensors, PO Box 1300, FI-33101 Tampere, Finland. martin.albers@vtt.fi
Munter
Tony
T
Laaksonen
Päivi
P
Vikholm-Lundin
Inger
I
D000178:Acrylamides; D017979:Gold Colloid; D006801:Humans; D007118:Immunoassay / Q000379:methods; D018796:Immunoconjugates / Q000737:chemistry*; D007074:Immunoglobulin G / Q000032:analysis*; D053758:Nanoparticles; D010316:Particle Size; D011108:Polymers / Q000737:chemistry*; D020349:Surface Plasmon Resonance; D008063:Thioctic Acid / Q000031:analogs & derivatives
D000178:Acrylamides; D017979:Gold Colloid; D018796:Immunoconjugates; D007074:Immunoglobulin G; D011108:Polymers; D008063:Thioctic Acid; C013091:lipoamide; C066443:N-methylolacrylamide