Immobilization of Protein Molecules onto Homogeneous and Mixed Carboxylate-Terminated Self-Assembled Monolayers

Abstract

The attachment of biomolecules, in particular proteins, onto solid supports is fundamental in the development of advanced biosensors, bioreactors, affinity chromatographic separation materials, and many diagnostic techniques. In addition, the effective investigation of biomolecular structure and function with scanning probe microscopy often requires a strong attachment of the biomolecule to a substrate. Here, we investigate the binding of the protein catalase to gold surfaces modified by self-assembled monolayers (SAMs). The chemical and physical adsorption of the protein molecules onto SAMs of 3-mercaptopropanoic acid (3-MPA), 11-mercaptoundecanoic acid (11-MUA), and a mixture of the two acid thiols (mixed) was investigated by utilizing tapping mode atomic force microscopy, scanning tunneling microscopy, surface plasmon resonance (SPR), static secondary ion mass spectrometry, and X-ray photoelectron spectroscopy. The surface concentration of catalase adsorbed on the SAMs decreased in the following order: mixed > 11-MUA > 3-MPA. Utilizing the terminal carboxylic acid functionalities, catalase was immobilized with a water-soluble carbodiimide and N-hydroxysuccinimide (NHS). Immobilization resulted in increased coverage of the protein. SPR studies on silver surfaces modified by these SAMs indicate that immobilization of carbodiimide and NHS decreased in the same order, namely mixed > 11-MUA > 3-MPA.

Insight into Serum Protein Interactions with Functionalized Magnetic Nanoparticles in Biological Media

“…36 Hence, although the net charge of a protein is negative, positively charged patches on the protein surface are also present due to the heterogeneous distribution on the protein surface. 37,38 Park et al demonstrated positive charge patches in bovine serum albumin (BSA) (that is otherwise negatively charged), 39 allowing BSA to bind with a strong polyanion, as also observed in this work and in a previous report. 32,38 The protein could also carry multiple positive charges and therefore be able to replace the counterions on a polymer surface.…”

Surface Plasmon Resonance Imaging of Biomolecular Interactions on a Grating-Based Sensor Array

“…Indeed, mixed SAMs containing long and short alkanethiols have proven most effective for protein immobilization. 47 A short adsorption time can also help minimize multilayer formation onto the gold surface. 56 SPR images of the grating arrays were acquired by collecting the reflected light intensity (R p /R s ) over the sample surface at a fixed angle of incidence and a single wavelength with the grating orientation perpendicular to the light path.…”

Wet chemical routes to the assembly of organic monolayers on silicon surfaces via the formation of Si–C bonds: surface preparation, passivation and functionalization

“…58,59,225 We reason this as the major cause behind the limited number of literature examples on this directive approach, while on the other hand the step-wise NHS/EDC activation scheme (Si-RCOOH to Si-RC(O)NHS) remain widespread in surface science. 50,51,226,227 It is also worth noting that an optimization study to evaluate the optimal range of NHS and EDC concentrations and molar ratios has only very recently been published for porous silicon substrates by Sam et. al.…”