AFM and MFM techniques for enzyme activity imaging and quantification

Abstract

In this work, a new approach for enzyme activity monitoring is suggested. It is based on the realtime imaging, by atomic force microscopy (AFM) and magnetic force microscopy (MFM), of the degradation of a nanoparticles-loaded enzyme responsive layer, namely Fe 3 O 4-nanoparticlesloaded gelatin. The roughness analysis allowed the quantification of the trypsin-induced ferrogel degradation and the correlation of the AFM and MFM data obtained.

Rapid disposable lipase activity sensor for automatic industrial application

“…The corresponding RSD values calculated from the obtained results are presented in Table 2. As seen there, all the obtained RSD values exceed the blank sample ones, because the sensitive layer enzymatic degradation by the lipase does not occur gradually but irregularly [24]. Some parts of the sensitive/dielectric layer become thinner, providing higher capacitance than others causing irregularity of the capacitance change during the hydrolysis process resulting in increased RSD, respectively.…”

“…mL À1 at 1.84% RSD determined by a series of 10 sensors. The sensitive layer irregular degradation [24] together with the nonlinearity of the capacitance versus the sensitive/dielectric layer thickness curve cause precision decrease in case of higher activities determination.…”

“…The irregular thickness of the sensitive layer (resulting from the applied deposition procedure: drop coating) combined with the irregular degradation of the sensitive layer during the measurement [24] is the second reason causing insufficient reproducibility of the sensor response not allowing its application automatically. This deposition technique has been chosen because it allows deposition of thin layers.…”

Bridging Biochemical Activities with Conformational Dynamics Observed in Atomic Force Microscopy

“…Additionally, once established, translocation activity can be used as a metric to quantitatively compare and to optimize supported lipid bilayer systems. Through the years there have been attempts to verify that surface-adsorbed proteins retain biochemical activity [9,[54][55][56][57][58][59][60][61][62]. Early in the field of biological AFM investigation, transient deflections of cantilevers positioned above surface adsorbed lysozymes were reported to result from enzymatic turn over, though following this strategy is challenging due to low frequency instrumental noise (i.e., tip-sample drift) [9].…”

Using steered molecular dynamic tension for assessing quality of computational protein structure models

“…AFM conventionally scans a sample by a probe and can measure the force between the probe and the sample surface or can be applied to image the sample shape 17 . With regard to proteins, AFM has been used to measure the binding force between a protein and a ligand, 18 to characterize enzyme activity, 19 to study unfolding pathways 20–22 and mechanical stability of proteins 23 . Early applications of SMD used to mimic AFM on biological systems were performed by the Schulten group to study the unbinding of the avidin‐biotin complex 24 and the unfolding pathway of titin IG domains, 25 which showed good agreement with contemporary experiments using AFM 26 .…”