Biomedical Engineering Reference
In-Depth Information
Table 4.1
Kinetics and afinity constants for the interactions of functionalised CNTs
with ODN-CpG motifs
NH3
+
Cl
-
O
O
O
NH
3
+
Cl
-
N
O
NH
O
N
Functionalised CNTs
Compound 2a: SWCNTs
Compound 2b: MWCNTs
NH
3
+
Cl
-
Compound 5: SWCNTs
k
on
[M
-1
s
-1
]
(5.19 ± 1.55)
*
10
5
(1.23 ± 0.14)
*
10
6
k
off
[s
-1
]
(2.99 ± 1.46)
*
10
-3
(4.85 ± 0.35)
*
10
-3
K
d
[M]
(5.88 ± 2.30)
*
10
-9
(3.99 ± 10.66)
*
10
-9
Subsequently, the immunostimulatory properties of the complexes were
tested
in vitro
.
24
In the irst series of experiments, both SWCNTs (
2a
and
5
) and MWCNTs
(
2b
) were selected. However, the MWCNTs with the free amino group
demonstrated much lower afinity for ODN-CpG sequences than did SWCNTs.
The difference could be attributable to both dimension and charge distribution
of the different CNTs. It was observed that SWCNT-Lys-NH
3
+
(compound
5
)
and MWCNT-NH
3
+
(compound
2b
) displayed the same amount of positive
charges. However, a stronger avidity effect could have been involved, resulting
from the closer proximity of the two ammonium groups in SWCNT-Lys-NH
3
+
than MWCNT-NH
3
+
and a consequent double electrostatic interaction for
ODN-CpG molecules. Therefore, SWCNTs represented the CNTs of choice for
the following experiments, in which various excess ratios of CNTs to a minimal
immunostimulatory dose of the ODN-CpG motifs were incubated with naïve
mouse splenocytes. In particular, complexes of SWNT-Lys-NH
3
+
at 18:1 and
9:1 ratios over ODN-CpG increased its immunostimulatory properties by
58% and 45%, respectively, while lower ratios did not show remarkable
results. This is most probably due to the fact that a high excess of positive
charges of CNTs
over ODN-CpG neutralised the negative charges of the DNA
sequences, thus minimising the repulsion effects with the negatively charged
cell membrane and promoting ODN-CpG's cellular internalisation.
4.4 IMMUNOGENICITY OF CARBON NANOTUBES
As reported in previous paragraphs, CNTs have been shown to be
advantageous as antigen-presenting systems. However, it is still necessary
to fully understand the role of functionalised CNTs in the immune response
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