Labelling of colloidal gold with protein. A quantitative study using beta-lactoglobulin

Area of Science:

• Bioconjugation Chemistry
• Materials Science
• Electron Microscopy

Background:

• Colloidal gold nanoparticles are widely used as labels in electron microscopy.
• Detailed understanding of macromolecule adsorption onto gold particles is limited.
• This limits optimization of gold marker preparation and performance.

Purpose of the Study:

• To quantitatively study the adsorption isotherm of proteins onto colloidal gold particles.
• To determine the adsorption capacity and stability of protein layers on gold.
• To assess the functional integrity of adsorbed proteins for antibody binding.

Main Methods:

• Quantitative adsorption isotherm studies using tritiated goat beta-lactoglobulin (β L).
• Chemical modification (iodination with 125I) to assess impact on adsorption.
• Ellipsometry on nickel-coated slides to study adsorption on metallic surfaces.
• Antibody binding assays to evaluate protein functionality post-adsorption.

Main Results:

• Adsorption followed a Langmuir-type monolayer model.
• A maximum of 13-14 β L molecules adsorbed per 12 nm gold particle, forming a compact monolayer.
• Adsorbed β L molecules retained their capacity to bind anti-β L antibodies.
• Adsorption was irreversible, and adsorbed proteins could not be displaced by polyethylene glycol.
• Optimal antibody binding required >5 β L molecules per particle.

Conclusions:

• Protein adsorption onto colloidal gold forms stable, compact monolayers.
• The number of adsorbed molecules influences the marker's ability to bind antibodies.
• This quantitative data aids in the rational design of improved colloidal gold-based electron microscopy labels.