Assessment of Small Cellular Particles from Four Different Natural Sources and Liposomes by Interferometric Light Microscopy

Abstract

Small particles in natural sources are a subject of interest for their potential role in intercellular, inter-organism, and inter-species interactions, but their harvesting and assessment present a challenge due to their small size and transient identity. We applied a recently developed interferometric light microscopy (ILM) to assess the number density and hydrodynamic radius (Rh) of isolated small cellular particles (SCPs) from blood preparations (plasma and washed erythrocytes) (B), spruce needle homogenate (S), suspension of flagellae of microalgae Tetraselmis chuii (T), conditioned culture media of microalgae Phaeodactylum tricornutum (P), and liposomes (L). The aliquots were also assessed by flow cytometry (FCM), dynamic light scattering (DLS), ultraviolet-visible spectrometry (UV-vis), and imaging by cryogenic transmission electron microscopy (cryo-TEM). In Rh, ILM showed agreement with DLS within the measurement error in 10 out of 13 samples and was the only method used here that yielded particle density. Cryo-TEM revealed that representative SCPs from Tetraselmis chuii flagella (T) did not have a globular shape, so the interpretation by Rh of the batch methods was biased. Cryo-TEM showed the presence of thin filaments in isolates from Phaeodactylum tricornutum conditioned culture media (P), which provides an explanation for the considerably larger Rh obtained by batch methods than the sizes of particles observed by cryo-TEM images. ILM proved convenient for assessment of number density and Rh of SCPs in blood preparations (e.g., plasma); therefore, its use in population and clinical studies is indicated.

Autologous Platelet and Extracellular Vesicle-Rich Plasma as Therapeutic Fluid: A Review

“…The quantity that reflects the size—the hydrodynamic radius R h —can be determined by dynamic light scattering (DLS) and interferometric light microscopy (ILM). In the isolate from plasma, the average R h assessed by DLS and ILM was 39 nm and 68 nm, respectively [ 47 ], indicating the presence of a large proportion of lipoproteins and protein complexes in the samples. The number density of EVs in plasma as determined by ILM was about 10 11 /mL [ 47 ].…”

“…In the isolate from plasma, the average R h assessed by DLS and ILM was 39 nm and 68 nm, respectively [ 47 ], indicating the presence of a large proportion of lipoproteins and protein complexes in the samples. The number density of EVs in plasma as determined by ILM was about 10 11 /mL [ 47 ].…”

“…In the recently developed method that is referred to as Interference Light Microscopy (ILM), the sample is illuminated by a LED light [ 47 ]. Interference enhances the information on the scattered light.…”

“…The sample shown in Panel ( B ) was prepared as described in [ 4 ]. The samples shown in Panel ( C , D ) were prepared as described in [ 47 ]. …”

Chemistry towards Biology

“…Masar et al [ 26 ] focused on the evaluation of biologically active compounds using capillary electrophoresis, while the optimization of chromatographic techniques and conditions for the characterization of bioactive molecules, or for metabolomics and lipidomics, are discussed in the contributions of Szucs et al [ 27 ] and Cajka et al [ 28 ]. An interferometric light microscopy methodology for evaluating small cellular particles and liposomes was developed by Romolo et al [ 29 ]. Physical and analytical measurements of the difference between solid–liquid interfacial adsorption of proteins in their native and amyloid forms were carried out by Abraham et al [ 30 ].…”