Nanoscale roughness affects the activity of enzymes adsorbed on cluster-assembled titania films.

In this study, we investigated how the adsorption properties governed by the nanometer-scale surface morphology of cluster-assembled titanium oxide films influence the catalytic activity of immobilized serine-protease trypsin. We developed an activity assay for the parallel detection of physisorbed enzyme activity and mass density of the adsorbed proteins in microarray format. The method combines a microarray-based technique and advanced quantitative confocal microscopy approaches based on fluorescent labeling of enzymes and covalent labeling of active sites of surface-bound enzymes. The observed diminishing trypsin binding affinity with increasing roughness, as opposed to the steep rise in its saturation uptake, was interpreted as heterogeneous nucleation-driven adsorption of trypsin at the rough nanoporous titania surface. The increase in relative activity of adsorbed trypsin is proportional to the fractional saturation of titania surfaces, expressed as percentage of saturation uptake. In turn, the specific activity, that is, the ratio of active proteins to the absolute number of adsorbed proteins, drops with growing saturation uptake and surface roughness, witnessing a reduction in the accessibility of enzyme active sites. Both geometrical constraints of titania nanopores and the clusterwise adsorption of trypsin were identified as the key factors underpinning the steric hindrance of the immobilized enzyme. These findings are relevant for the optimization of rough nanoporous surfaces as carriers of immobilized enzymes. The proposed activity assay is particularly advantageous in the screening of candidate materials for enzyme immobilization.

Simultaneous recording of skin blood pulsations at different vascular depths by multiwavelength photoplethysmography.

A new technique for parallel recording of reflection photoplethysmography (PPG) signals in a broad spectral band (violet to near-infrared) has been developed, and its potential for assessment of blood microcirculation at various depths from the skin surface is discussed. PPG signals have been simultaneously detected at cw laser wavelength sets comprising 405, 532, 645, 807, and 1064 nm. Various signal baseline responses to breath holding and different shapes of the PPG pulses originated from the same heartbeat but recorded at different wavelengths have been observed, indicating a depth variety of the skin blood pulsation dynamics.