Abstract
The present work addresses some fundamental aspects in the preparation of protein-conjugated gold nanoparticles, in order to ensure an appropriate final product. Ten broadly available and/or easy to implement analytical tools were benchmarked and compared in their capacity to provide reliable and conclusive information for each step of the procedure. These techniques included transmission electron microscopy, UV/VIS spectroscopy, dynamic light scattering, zeta-potential, Fourier-transformed infrared spectroscopy, colloidal stability titration, end-point colloidal stability analysis, cyclic voltammetry, agarose gel electrophoresis and size-exclusion chromatography (SEC). Four different proteins widely used as adaptors or blocking agents were tested, together with 13 nm gold nanoparticles containing different surface chemistries. Among all tested techniques, some of the least popular among nanomaterial scientists probed to be the most informative, including colloidal stability, gel electrophoresis and SEC; the latter being also an efficient purification procedure. These three techniques provide low-cost, low time consuming, sensitive and robust ways to assess the success of the nanoparticle bioconjugation steps, especially when used in adequate combinations.
Highlights
Protein attachment to gold nanoparticles (AuNPs) is a very common procedure in diverse scientific fields [1, 2, 3] and is no longer limited to research laboratories with experience in nanochemistry
A combination of techniques needs to be taken together to obtain a correct interpretation of the physical phenomena involved in each step of the bioconjugation process
We have provided a critical evaluation of ten different techniques and their utility to inform about different stages of the process
Summary
Protein attachment to gold nanoparticles (AuNPs) is a very common procedure in diverse scientific fields [1, 2, 3] and is no longer limited to research laboratories with experience in nanochemistry. Despite this being a desired outcome of decades of research in material sciences, the widespread use of protein-AuNP conjugates comes at a cost. To achieve the goal of efficient protein binding to AuNPs, different strategies have been employed These strategies can involve both covalent immobilization (typically by carbodiimide chemistry) and non-covalent adsorption [3, 6, 7, 8]
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