Obtaining optimization and application of multifunctional conjugates of gold nanoparticles with antibodies and peroxidase for the detection of bacterial lipopolysaccharides

Abstract

Multifunctional nanoparticles containing antibodies are promising probes for highly sensitive detection of various antigens. Gold nanoparticles (GNPs) themselves have a plasmon resonance and are able to signal the interaction of antibodies from conjugates with an antigen. The presence of one or more enzyme molecules on the surface of a GNP can increase the resolution of detection methods based on nanoparticles by hundreds or thousands of times.
To obtain multifunctional nanoparticles, solutions of GNPs (d = 15 nm; 1012 particles/ml) were mixed with 100 µl of peroxidase solution (20 µg/ml – ½ of the “golden number”). After 15 minutes, 100 μl of an antibody solution to the O-antigen of Azospirillum baldaniorum Sp245 (C = 50 μg/ml) was added. Conjugates were precipitated by centrifugation (15,000 g; 30 min) and redissolved in water. The resulting conjugates exhibited peroxidase activity and specifically interacted with the O antigen of A. baldaniorum Sp245 in dot analysis.
Comparison of the enzymatic activity of the solutions from different stages of conjugation showed that about 95% of the enzyme molecules are adsorbed on the surface of the nanoparticles. The activity of the enzyme in the composition of conjugates is 16% lower than that of solved peroxidase molecules. Calculations showed that 1 nanoparticle of the conjugate contains, on average, 17 peroxidase molecules. Therefore, a single antigen-antibody interaction will be accompanied by a 14-fold increase in enzymatic activity. The resulting multifunctional GNPs can be used as a variant of the sandwich method for more sensitive detection of bacteria and their surface antigens.


File with abstract

Speaker

Alina Matora, Anastasia Astankova
SGU
Russia

Discussion

Ask question