Multicompartment Lipid Nanocarriers for Targeting of Cells Expressing Brain Receptors - Self-Assembled Supramolecular Architectures: Lyotropic Liquid Crystals - page 338

0.005

1

0.001
5×10 −4

2

1×10 −4

5×10 −5

1×10 −5

0.01 0.02 0.05

Log(q) [Å −1 ]

0.10 0.20

Figure 11.9 Small - angle X-ray scattering (SAXS) curves of diluted lipid nanocarrier
systems: MO/DOPE - PEG 2000 (98/2 mol %) (curve 1) and MO/DOPE-PEG 2000 /EPA
(83/2/15 mol %) (curve 2).

following the SH-SY5Y cell differentiation by treatment with retinoic acid,
was evidenced by confocal fl uorescence microscopy (for the employed fl uo-
rescently labeled antibody,

650 nm). The images were
taken from SH-SY5Y cells treated and fi xed on glass slides in order to visual-
ize the expression of TrkB receptors by detecting the fl uorescence of a second-
ary antibody that recognizes the anti-TrkB antibody, which was bound to the
neurotrophin receptor TrkB. Controls were acquired in order to ensure the
nonfl uorescent character of the cells themselves and to check that the employed
primary antibody was specifi c to the TrkB receptor. The untreated SH-SY5Y
cells had no autofl uorescence at the studied excitation wavelength (Fig. 11.10a).
No fl uorescence was detected when the cells were treated by retinoic acid and
when the secondary antibody was incubated with the cells in the absence of a
primary antibody (anti-TrkB). These results confi rmed the antibodies specifi c-
ity and the nonfl uorescence of the cells treated with retinoic acid in the
absence of a primary antibody (Fig. 11.10b). The unusual morphology of the
fi xed cells was explained by the drastic shock employed for their fi xation on
microscopic slides.
The expression of the TrkB receptors was established by superimposing the
fl uorescence images of the SH-SY5Y cells with those recorded in a Nomarski
mode. The fl uorescently labeled secondary antibody was proven to be specifi c
to the primary anti-TrkB antibody as it did not bind to the cells in the lack of
a primary antibody (that recognizes the expressed neurotrophin receptor). The
confocal fl uorescence images of cells, treated for 4 days with retinoic acid
(10

λ ex = 633 nm and λ em >

M) are shown in Figure 11.11. The cells exhibited fl uorescence due to the
Alexa Fluor 633 anti-IgG antibody, which binds to the antihuman TrkB mouse

μ