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Microscopic validation of macroscopic in vivo images enabled by same-slide optical and nuclear fusion.
[lymphangioleiomyomatosis]
It
is
currently
difficult
to
determine
the
molecular
and
cellular
basis
for
radioscintigraphic
signals
obtained
during
macroscopic
in
vivo
imaging
.
The
field
is
in
need
of
technology
that
helps
bridge
the
macroscopic
and
microscopic
regimes
.
To
solve
this
problem
,
we
developed
a
fiducial
marker
(
FM
)
simultaneously
compatible
with
2
-
color
near-infrared
(
NIR
)
fluorescence
(
700
and
800
nm
)
,
autoradiography
,
and
conventional
hematoxylin-eosin
(
HE
)
histology
.
T
he
FM
was
constructed
from
an
optimized
concentration
of
commercially
available
human
serum
albumin
,
700
-
and
800
-
nm
NIR
fluorophores
,
(
99
m
)
Tc-pertechnetate
,
dimethyl
sulfoxide
,
and
glutaraldehyde
.
Lymphangioleiomyomatosis
cells
coexpressing
the
sodium
iodide
symporter
and
green
fluorescent
protein
were
labeled
with
700
-
nm
fluorophore
and
(
99
m
)
Tc-pertechnatate
and
then
administered
intratracheally
into
CD-
1
mice
.
After
in
vivo
SPECT
imaging
and
ex
vivo
SPECT
and
NIR
fluorescence
imaging
of
the
lungs
,
30
-
μm
frozen
sections
were
prepared
and
processed
for
800
-
nm
NIR
fluorophore
costaining
,
autoradiography
,
and
HE
staining
on
the
same
slide
using
the
FMs
to
coregister
all
datasets
.
Optimized
FMs
,
composed
of
100
μM
unlabeled
human
serum
albumin
,
1
μM
NIR
fluorescent
human
serum
albumin
,
15
%
dimethyl
sulfoxide
,
and
3
%
glutaraldehyde
in
phosphate-buffered
saline
(
pH
7
.
4
)
,
were
prepared
within
15
min
,
displayed
homogeneity
and
stability
,
and
were
visible
by
all
imaging
modalities
,
including
HE
staining
.
Using
these
FMs
,
tissue
displaying
high
signal
by
SPECT
could
be
dissected
and
analyzed
on
the
same
slide
and
at
the
microscopic
level
for
700
-
nm
NIR
fluorescence
,
800
-
nm
NIR
fluorescence
,
autoradiography
,
and
HE
histopathologic
staining
.
W
hen
multimodal
FMs
are
combined
with
a
new
technique
for
simultaneous
same-slide
NIR
fluorescence
imaging
,
autoradiography
,
and
HE
staining
,
macroscopic
in
vivo
images
can
now
be
studied
unambiguously
at
the
microscopic
level
.
Diseases
Validation
Diseases presenting
"unlabeled human serum albumin"
symptom
lymphangioleiomyomatosis
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