Proteins immobilized on nanoparticles – determination of the surface coverage
Jaroslaw Grobelny
University of Lodz
Dr Jaroslaw Grobelny Assoc. Prof. finished his doctoral degree in physics in Lodz University, Poland 2003. From 2004 he spent 3 years at the National Institute of Standers and Technology, Maryland, USA. In 2010 he has completed his habilitation in materials science. He has published more than 60 paper in international peer reviewed journals. His research interest is focus on nanoparticles synthesis, characterization and surface modification.
Abstract
Introduction Proteins can be immobilised on the surface of nanoparticles (NPs) by two main roads: covalent bounding or via adsorption process. Regardless the method, the immobilisation of proteins on NPs allows to... [ view full abstract ]
Introduction
Proteins can be immobilised on the surface of nanoparticles (NPs) by two main roads: covalent bounding or via adsorption process. Regardless the method, the immobilisation of proteins on NPs allows to design a protein-particle hybrid system which may exhibit a new properties that result from synergistic effect of combination of its components (NPs and protein). Hence, the precise characterization of NPs and proteins, as well as the determination of the amount of proteins on NPs, is crucial. There are several methods available to characterize NPs and proteins separately, but the quantification of the amount of protein on the single NP surface is still challenging. In this work a simple, reproducible and highly sensitive analytical method to determine the amount of protein adsorbed on a single NP will be presented.
Methods
Metallic nanoparticles (mNPs) synthesized via chemical reduction method were modified with proteins (catalase-CAT) by incubation. Dynamic Light Scattering was used to monitor the agglomeration state of colloids before and after the immobilisation of CAT as well as to investigate the hydrodynamic size of mNPs modified with CAT. Morphological studies of the mNPs before and after modification were performed by S-TEM. Determination of CAT adsorption was carried out with different electrophoresis protocols to find the optimal conditions for quantification of the surface coverage of mNPs by catalase.
Results
It was found that the modification process with CAT did not disturb the stability of NPs. Particles were colloidally stable after the modification without any aggregates or agglomerates. The hydrodynamic size of protein-modified ANPs increased compared to non-modified which confirmed the successful modification process. The determination of the surface coverage of NPs with CAT was performed based on the native-PAGE for which the most sensitive conditions were found.
Discussion
The presented method allowed for the identification and quantification of the amount of CAT adsorbed on the surface of NPs in a colloidal state. The obtained results prove that the method is effective and versatile and can be successfully used for the identification and quantification of proteins adsorbed on the surface of different types of colloidal NPs.
NSC of Poland 2013/09/B/NZ7/01019
Authors
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Jaroslaw Grobelny
(University of Lodz)
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Katarzyna Ranoszek-Soliwoda
(University of Lodz)
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Ewa Czechowska
(University of Lodz)
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Emilia Tomaszewska
(University of Lodz)
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Grzegorz Celichowski
(University of Lodz)
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Janusz Szemraj
(Medical University of Lodz)
Topic Area
Targeted drug delivery and nanocarriers
Session
OS2a-2 » Nanomedicine and nanobiotechnology (14:30 - Thursday, 19th October, Room 2)
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