Biomedical Engineering Reference
In-Depth Information
molecular profiling of cancer markers (Her2/
neu
, EGFR, and EpCAM) on human
cells was demonstrated with CLIO nanoparticles directly conjugated to monoclonal
antibodies. With the development of the second-generation NMR-2 and the
highly magnetic MnFe
2
O
4
nanoparticles, subsequent cellular detection sensitivity
was remarkably improved to approximately single-cell level, far surpassing the
sensitivity of other conventional clinical methods (Fig.
9.10
a). Furthermore, a new
assay protocol was established that reports the expression level of a specific
biomarker and the target cell density.
To independently measure cell numbers by NMR, we exploited a phenomenon
of low-grade phagocytosis of nontargeted MNPs by tumor cells [
59
]. When mam-
malian cells were incubated (15 min at 37
ı
C) with unmodified MNPs (MNP-ˆ),
linear and cell-number-dependent R
2
changes (R
2
) were observed (Fig.
9.10
b).
Interestingly, these changes were similar across a wide variety of cell types. The
results were fitted to Eq.
9.2
; R
2
n
C
,wherer
2
is the cellular relaxivity
for MNP-ˆ and n
C
is the cell concentration (N
C
=V). The cellular relaxivities (r
2
)
were statistically identical (p > 0:99) among different tumor cell lines, suggesting
that the method may provide a universal measure for estimating n
C
.
Using Eq.
9.2
and the cell density information (above), the expression level ()
of a select marker was defined as
r
2
D
R
Ab
2
r
Ab
2
r
Ab
2
r
2
n
C
Ab
D
R
2
D
n
C
D
;
(9.4)
r
2
where R
Ab
2
and r
Ab
2
are R
2
changes and the cellular relaxivity, respectively,
with a marker-specific MNP. In this normalized form, now reports the
cellular
expression level of a targeted marker, providing a way to molecularly profile target
cells regardless of cell numbers in a sample. The method was extensively verified
by comparing NMR measurements to other standard methods (flow cytometry,
Western blotting) [
22
,
27
,
28
]. In one set of experiments, we measured the expres-
sion level of HER2 in breast cancer cell lines (Fig.
9.10
c). The measured
HER
2
.
10
3
cells) showed good agreement
(R
2
>98%) with both flow cytometry (requiring
R
HER
2
2
=R
2
/ from NMR (requiring
D
10
5
cells) and Western blotting
10
7
cells), validating the analytical capability of DMR. Note that DMR
detection was much faster (
(requiring
15 min) and performed using >10
2
times fewer cells.
9.6.2
Clinical Trial
Through the integration of the complementary DMR and BOND technologies, this
chip-based NMR detection platform has been applied in clinical trials of cancer
cell profiling [
26
]. A total of 50 patients with suspected abdominal malignancies
were enrolled. Each patient underwent fine-needle aspiration (FNA) using a 22-G
needle, followed by routine core biopsies (17-G needle) for conventional analysis.
The FNA samples were aliquoted and profiled for 11 predefined cellular markers:
