Nanoparticles for Drug and Gene Delivery in Treating Diseases of the Eye - Ocular Pharmacology and Toxicology - page 294

Particle size is an important parameter that not only affects the
degradation of the polymer matrix but also influences the disposi-
tion of the particles once they have been injected into the eye
[ 21 , 22 ]. There is no single standard technique for measuring the
particle size of nanoparticles. A variety of methods like dynamic
light scattering (DLS), scanning electron microscopy (SEM), trans-
mission electron microscopy (TEM), and nanoparticle tracking
analysis (NTA) can be employed for determining the particle size
[ 7 , 16 , 17 , 23 , 24 ]. DLS, which is one of the most commonly used
methods for measuring particle size is discussed here. DLS is a
characterization technique that reports the particle size in terms
of the hydrodynamic diameter of the particles [ 25 ]. It is advanta-
geous because it is non-destructive, making sample recovery possi-
ble, uses a small amount of sample and is quick and easy to perform
[ 26 ]. Step-by-step procedure for measuring particles size by DLS is
as follows:

3.2 Characterization
of Nanoparticles

3.2.1 Particle Size

1. 1 mg of PLA/PLGA nanoparticles are weighed in an eppen-
dorf tube.
2. 1 ml filtered, de-ionized water is added to the particles ( see
Notes 29 and 30 ).
3. The particles are suspended in water by gently shaking the
eppendorf tube ( see Note 31 ).
4. The dispersion prepared in step 3 can be analyzed using the
Zetasizer Nano ZS (Malvern ® Instruments) ( see Note 32 ).

Particle geometry and surface features of the nanoparticles can be
studied using SEM and TEM. The detailed procedure for the two
methods is as follows.

3.2.2 Morphology

1. The particles are mounted on aluminum stubs.
2. Following mounting, the particles are coated with a layer of
gold using a sputter coater (Anatech, CA, USA).
3. After coating, the particles are examined under an electron
microscope set at 5-10 kV [ 16 , 17 ]( see Note 33 ).

Scanning Electron
Microscopy (SEM)

1. A droplet of the nanoparticle suspension is placed on a flexible
plastic film (Parafilm ® )( see Note 34 ).
2. Carbon-coated grid is floated on the droplet of nanoparticle
suspension to permit the adsorption of the nanoparticles onto
the grid.
3. The grid is blotted with a filter paper and air dried for 5 min
( see Note 35 ).
4. After drying, the grid is transferred onto a drop of uranyl
acetate or ammonium molybdate.

Transmission Electron
Microscopy (TEM)