Environmental Engineering Reference
In-Depth Information
nanoparticles. Thus the surface hydrophobicity and the functional groups present
affect the protein adsorption patterns.
Nanoparticles of cross-linked copolymers of NIPAM and BAM (70-700 nm)
and two comonomer ratios 50:50 and 85:15 were evaluated to study the effects of
nanoparticle curvature and hydrophobicity on the nature and identity of adsorbed
plasma proteins [
65
]. The hydrophobicity of the particle surface influenced both the
amount and identity of the proteins bound to the particles [
66
]. The less hydropho-
bic 85:15 NIPAM-BAM copolymer particles showed virtually no protein adsorbed
on them (except some HSA), in strong contrast to the more hydrophobic 50:50
particles which showed high levels of protein adsorption. The amount of bound
protein varied with size, and scaled with the amount of available surface area.
However, the protein pattern was the same for all sizes and apolipoprotein AI was
always the most abundant protein recovered. This indicated that the surface curva-
ture was not a major determining factor for the relative affinities of proteins for the
particles.
15.2.7
Incubation Medium
The incubation medium for the particles can be blood, plasma or serum. Ideally,
blood should be used, but it is very difficult to separate the nanoparticles from the
blood cells. The monocytes present in the blood start to phagocytose the particles,
thus the particles disappear. In vivo, this happens only to a limited extent, because
particles are taken up, e.g. within 5 min by the liver macrophages or accumulate
somewhere else. Therefore, in general, plasma or serum is used instead of blood.
However, serum and plasma yield different patterns, because, e.g. serum is
depleted from proteins of the complement system due to the coagulation process
(e.g. lack of C3 and fibrinogen). The use of plasma as an incubation medium for the
in vitro evaluation of colloidal drug carriers is widely reported [
67
,
68
]. However
the plasma contains compounds to inhibit coagulation, e.g. citrate or heparin, which
can also interfere with the protein adsorption process. To minimise such effects and
to obtain comparable results, macromolecular heparins should be avoided. Hence,
citrate plasma is often used as an incubation medium.
15.2.8 Ratio of Sample to the Incubation Medium
The ratio of particle suspension to incubation medium is a very crucial parameter.
In case the ratio is too high (too many particles, too large of a surface area), some
high affinity and lowly concentrated proteins in the plasma or serum might be
depleted. They would have been adsorbed in a higher amount if more protein would
have been available. The protocol “NCL Method ITA-4” [
69
] suggests to mix 1 mL
plasma with 1 mL particle suspension (concentration 1 mg/mL
¼
0.1 %).
