Fluorescence microspectroscopy as a tool to study mechanism of nanoparticles delivery into living cancer cells

Abstract

Lack of better understanding of nanoparticles targeted delivery into cancer cells calls for advanced optical microscopy methodologies. Here we present a development of fluorescence microspectroscopy (spectral imaging) based on a white light spinning disk confocal microscope with emission wavelength selection by a liquid crystal tunable filter. Spectral contrasting of images was used to localize polymer nanoparticles and cell membranes labeled with fluorophores that have substantially overlapping spectra. In addition, fluorescence microspectroscopy enabled spatiallyresolved detection of small but significant effects of local molecular environment on the properties of environment-sensitive fluorescent probe. The observed spectral shift suggests that the delivery of suitably composed cancerostatic alkylphospholipid nanoparticles into living cancer cells might rely on the fusion with plasma cell membrane.

Lysolipid containing liposomes for transendothelial drug delivery

“…Since biological membranes are composed of dynamically condensed domains surrounded by fluid domains, it has been suggested that, above the CMC, alkyl phospholipids can insert into both kinds of phases: as monomers into the condensed phase and as a group of monomers into the fluid phase [32]. This is also in agreement with fluorescence microspectroscopy data, which show that lipophilic phospholipid fluorescent probe NBD-PC, where 7-nitrobenz-2-oxa-1,3-diazol-4-yl (NBD) is attached to a phosphatidylcholine phospholipid is immediately transferred into cells after addition of liposomes with higher than 1:1 ratio of perifosine to cholesterol, whereas liposomes with lower ratio do not interact with cells [26,33]. With electron paramagnetic resonance spectroscopy it was similarly shown that hydrophilic spin probe encapsulated in liposomes with lower than 1:1 ratio of perifosine to cholesterol does not enter cells, whereas starts entering cells immediately after incubation when it is encapsulated in liposomes with higher than 1:1 ratio of perifosine to cholesterol [34].…”

Transient submicron temperature imaging based on the fluorescence emission in an Er/Yb co-doped glass–ceramic

“…1. The system was built on an inverted microscope Nikon Eclipse TE 2000-E platform with a combination of a fluorescence unit CARV II spinning disk confocal module (BD Biosciences, Franklin Lakes, NJ, USA) that housed the excitation, dichroic and emission filters (BrightLine filters from Semrock, Rochester, NY, USA) as well as a multiple-hole spinning disk for the confocal imaging [33]. Objective lenses with 10× and 40× magnification were used in these experiments to provide the high magnification and the desired high spatial resolution.…”

Spectral selective fluorescence molecular imaging with volume holographic imaging system

“…However, both¯lter technologies su®er from the polarization-dependent property, which leads to poor light transmission e±ciency (approximately 40%) and the additional drivers make the imaging system more complex and expensive. 8,9 As a newly developed spectral selecting technique, TFTF has sharp edge cut-o®s and high out-of-band optical densities. But TFTF needs an array of thin¯lm tunable¯lters to cover a wide wavelength range, because each single TFTF has a limited bandwidth.…”