Quartz Crystal Microbalance (QCM): Useful for Developing Procedures for Immobilization of Proteins on Solid Surfaces

Abstract

We demonstrate the combined use of liquid and air measurements with the quartz crystal microbalance (QCM) for quantitative analysis of multistep reaction procedures leading to immobilized proteins on solid surfaces. Reactions are conducted on the surfaces of QCM sensor crystals and are quantified by measurements of resonant frequency of the crystals before and after each reaction step. When reactions are conducted in the flow cell of the QCM in the presence of solvent, measurement of resonant frequency can be made in situ (liquid measurement). When reactions cannot be conducted in the flow cell because of temperatures or solvents not tolerated by the cell, frequency can be measured after evaporation of solvent (air measurement). Each reaction step can be analyzed by either liquid or air measurement so that the whole multistep procedure is addressed, no matter how diverse the chemical nature of the steps. We conducted identical multistep procedures on two different starting surfaces, gold and silica, and found comparable results.

Displacement-type Quartz Crystal Microbalance Immunosensing Platform for Ultrasensitive Monitoring of Small Molecular Toxins

“…Related to transducers, QCM has become an analytical tool for investigation of biomolecular interaction and clinical bioassay owing to its high sensitivity, low cost, real-time output, and label- or radiation-free entities . The QCM sensors measure the resonant frequency ( f ) by the standard oscillator technique. , The frequency shift (Δ f ) is usually explained by the Sauerbrey equation, which states that the frequency shift is linearly proportional to the change of surface mass (Δ m ) on the crystal. , Salmain et al designed a label-free QCM immunosensor for detection of staphylococcal enterotoxin A in milk based on the direct antigen–antibody reaction.…”

Antifouling Coatings Generated from Unsymmetrical Partially Fluorinated Spiroalkanedithiols

“…Together with ellipsometry, a common and easily accessible method, we chose to explore the use of other, more sensitive biophysical techniques, such as SPR − and QCM, − to examine the protein resistance of thin films derived from the adsorbates, as shown in Figure . The biophysical methods are needed not only to evaluate the validity of the ellipsometry data but also to gain broader insight into the antifouling performance of the unique “conflicted” interfaces prepared herein.…”

Continuous Monitoring of Enzymatic Reactions on Surfaces by Real-Time Flow Cytometry: Sortase A Catalyzed Protein Immobilization as a Case Study

“…Examples include the immobilization of fluorescent proteins on polymer microbeads and liposomes, 15−18 human and bacterial glycosyltransferases on sepharose particles, 19 single-chain variable antibody fragments on PEGylated polymer capsules, 20 fibronectin binding protein on CM5 biosensor chips, 21 and recombinant human thrombomodulin on glass slides. 22 Due to the limited number of techniques that enable continuous monitoring of protein attachment on surfaces (e.g., quartz crystal microbalance (QCM) 23,24 and surface plasmon resonance spectroscopy (SPR) 25,26 ), the efficiency of immobilization reactions is usually judged by end point assays evaluating functional properties of the obtained protein hybrid material, such as fluorescence intensity, enzymatic activity, or binding affinity. 12,13,15−22 This generally complicates the optimization of enzyme-catalyzed immobilization reactions and leads to the fact that process parameters such as concentration of enzyme and reactants, buffer conditions, temperature, and reaction time are in most cases chosen empirically.…”

“…Due to the limited number of techniques that enable continuous monitoring of protein attachment on surfaces (e.g., quartz crystal microbalance (QCM) , and surface plasmon resonance spectroscopy (SPR) , ), the efficiency of immobilization reactions is usually judged by end point assays evaluating functional properties of the obtained protein hybrid material, such as fluorescence intensity, enzymatic activity, or binding affinity. ,,− This generally complicates the optimization of enzyme-catalyzed immobilization reactions and leads to the fact that process parameters such as concentration of enzyme and reactants, buffer conditions, temperature, and reaction time are in most cases chosen empirically. In the case of immobilization reactions catalyzed by SrtA, another important, yet mostly neglected factor, is the choice of the enzyme variant employed for the particular coupling reaction.…”