Nano-domains are sub-light-diffraction-sized heterogeneous areas in the plasma membrane of cells, which are involved in cell signalling and membrane trafficking. Throughout the last thirty years, these nano-domains have been researched extensively and have been the subject of multiple theories and models: the lipid raft theory, the fence model, and the protein oligomerization theory. Strong evidence exists for all of these, and consequently they were combined into a hierarchal model. Measurements of protein and lipid diffusion coefficients and patterns have been instrumental in plasma membrane research and by extension in nano-domain research. This has led to the development of multiple methodologies that can measure diffusion and confinement parameters including single particle tracking, fluorescence correlation spectroscopy, image correlation spectroscopy and fluorescence recovery after photobleaching. Here we review the performance and strengths of these methods in the context of their use in identification and characterization of plasma membrane nano-domains.

Ammonia (NH3) emission from agriculture is an environmental threat and a loss of nitrogen for crop production. Mineral fertilizers and manure are significant sources of NH3; therefore, abatement technologies have been introduced to mitigate these emissions. The aim of this study was to demonstrate that low-cost measuring techniques are suitable to assess NH3 emissions in smaller plots, appropriate to test different managements. Two experiments were established to quantify NH3 emissions from urea application in a multi-plot design with radii of 5 (R5) and 20 m (R20). Field was bare soil partially surrounded by shelterbelts. Measurement techniques included passive flux samplers (Leuning), and passive concentration samplers (ALPHA) coupled to WindTrax dispersion model. NH3 emission from R5 was consistent with the emission from R20 when the surface-to-atmosphere exchange was not affected by shelterbelts, and wind speed near surface was greater than 1 m s−1. Both measurement methods gave unreliable NH3 quantification in combination with wind speed lower than 1 m s−1 and low emission strength. Cumulative emission over 60 h was 2% of the supplied N from the plots not affected by the shelterbelt, and 1% from the plots affected by shelterbelts, indicating that these structures can significantly reduce NH3 emissions.

In this study, we explore the use of line FRAP to detect diffusion in synthetic lipid membranes. The study of the dynamics of these membrane lipids can, however, be challenging. The diffusion in two different synthetic membranes consisting of the lipid mixtures 1:1 DOPC:DPPC and 2:2:1 DOPC:DPPC:Cholesterol was studied with line FRAP. A correlation between diffusion coefficient and temperature was found to be dependent on the morphology of the membrane. We suggest line FRAP as a promising accessible and simple technique to study diffusion in plasma membranes.

The formation of nanodomains in the plasma membrane are thought to be part of membrane proteins regulation and signaling. Plasma membrane proteins are often investigated by analyzing the lateral mobility. k-space ICS (kICS) is a powerful image correlation spectroscopy (ICS) technique and a valuable supplement to fluorescence correlation spectroscopy (FCS). Here, we study the diffusion of aquaporin-9 (AQP9) in the plasma membrane, and the effect of different membrane and cytoskeleton affecting drugs, and therefore nanodomain perturbing, using kICS. We measured the diffusion coefficient of AQP9 after addition of these drugs using live cell Total Internal Reflection Fluorescence imaging on HEK-293 cells. The actin polymerization inhibitors Cytochalasin D and Latrunculin A do not affect the diffusion coefficient of AQP9. Methyl-β-Cyclodextrin decreases GFP-AQP9 diffusion coefficient in the plasma membrane. Human epidermal growth factor led to an increase in the diffusion coefficient of AQP9. These findings led to the conclusion that kICS can be used to measure diffusion AQP9, and suggests that the AQP9 is not part of nanodomains.

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