88
Dissolve 3-[4,5-dimethylthiazole-2-yl]-2,5-diphenyltetrazolium
bromide (MTT) in PBS at a concentration of 1 mg/mL by using
vortexing or sonication and filtration. Store the solution at −20 °C
for 4 months and at −4 °C for few days. MTT is photosensitive so
use foil to protect it.
1. Dimethyl sulfoxide (DMSO).
2. Dulbecco’s phosphate-buffered saline (sterile PBS, 10 mM).
3. Sonicator.
4. Cellulose membrane dialysis bag of specific molecular weight
cutoff.
5. Magnetic hyperthermia apparatus.
6. EDTA-trypsin.
7. L-Poly lysine.
8. p-Formaldehyde.
9. Antiphotobleacher: b-Mercaptoethanol.
10. 96-Well plates, 6-well plates.
11. UV-vis microplate reader.
12. UV-vis spectrophotometer.
13. Glass coverslips.
14. Hematocytometer.
3 Methods
Tumoral tissues exhibit pH, thermal, and redox instabilities which
are taken into consideration when designing a polymeric nanocontainer (NCs) as a DDS. In more detail, cancer cells proliferate in a
more acidic environment; they are very sensitive to temperatures
between 40 and 45 °C and they tend to thrive in a hypoxic environment. These three basic characteristics can be exploited by fabricating a model triple-sensitive polymeric nanocontainer. For example,
as a temperature-sensitive polymer PNIPAm is commonly used and
has a lower critical solution temperature (LCST) of 32 °C where it
shows a reversible volume-phase transition. Another commonly
used polymer is methacrylic acid (MAA) which is pH sensitive and
under acidic environment it is fully protonated, facilitating the
release of an electrostatically bound drug and cellular uptake
through adsorptive endocytosis. Finally, polymers containing glutathione labile bonds like N,N′-(disulfanediylbis(ethane-2,1-diyl))
bis(2-methylacrylamide) (DSBMA) are employed in order to allow
site-specific cleavage in the reductive tumoral environment.
Nanocontainers synthesized with these kind of polymers can serve
as a model platform for intracellular release of anticancer drugs and
2.2.2 MTT Solution
Preparation
2.2.3 Other Reagents
and Materials
3.1 Drug Loading
and Release
Maria Theodosiou et al.
Dissolve 3-[4,5-dimethylthiazole-2-yl]-2,5-diphenyltetrazolium
bromide (MTT) in PBS at a concentration of 1 mg/mL by using
vortexing or sonication and filtration. Store the solution at −20 °C
for 4 months and at −4 °C for few days. MTT is photosensitive so
use foil to protect it.
1. Dimethyl sulfoxide (DMSO).
2. Dulbecco’s phosphate-buffered saline (sterile PBS, 10 mM).
3. Sonicator.
4. Cellulose membrane dialysis bag of specific molecular weight
cutoff.
5. Magnetic hyperthermia apparatus.
6. EDTA-trypsin.
7. L-Poly lysine.
8. p-Formaldehyde.
9. Antiphotobleacher: b-Mercaptoethanol.
10. 96-Well plates, 6-well plates.
11. UV-vis microplate reader.
12. UV-vis spectrophotometer.
13. Glass coverslips.
14. Hematocytometer.
3 Methods
Tumoral tissues exhibit pH, thermal, and redox instabilities which
are taken into consideration when designing a polymeric nanocontainer (NCs) as a DDS. In more detail, cancer cells proliferate in a
more acidic environment; they are very sensitive to temperatures
between 40 and 45 °C and they tend to thrive in a hypoxic environment. These three basic characteristics can be exploited by fabricating a model triple-sensitive polymeric nanocontainer. For example,
as a temperature-sensitive polymer PNIPAm is commonly used and
has a lower critical solution temperature (LCST) of 32 °C where it
shows a reversible volume-phase transition. Another commonly
used polymer is methacrylic acid (MAA) which is pH sensitive and
under acidic environment it is fully protonated, facilitating the
release of an electrostatically bound drug and cellular uptake
through adsorptive endocytosis. Finally, polymers containing glutathione labile bonds like N,N′-(disulfanediylbis(ethane-2,1-diyl))
bis(2-methylacrylamide) (DSBMA) are employed in order to allow
site-specific cleavage in the reductive tumoral environment.
Nanocontainers synthesized with these kind of polymers can serve
as a model platform for intracellular release of anticancer drugs and
2.2.2 MTT Solution
Preparation
2.2.3 Other Reagents
and Materials
3.1 Drug Loading
and Release
Maria Theodosiou et al.