Capillary Isoelectric Focusing of Proteins and Peptides Using an In-Line cIEF-ESI Interface with Improved MS Characteristics
Analytical Chemistry,
Год журнала:
2025,
Номер
97(1), С. 649 - 657
Опубликована: Янв. 1, 2025
Intact
protein
analysis
using
mass
spectrometry
(MS)
is
an
important
technique
to
characterize
and
provide
a
comprehensive
overview
of
complexity.
It
also
the
basis
"top-down"
approaches
in
proteomics
describe
proteoforms
single
protein's
post-translational
modifications
(PTMs).
MS-based
intact
proteins
benefits
from
high-resolution
separations
prior
electrospray
ionization.
Capillary
isoelectric
focusing
(cIEF)
separation
for
peptides
which
capable
separating
proteoforms.
MS
detection
coupled
cIEF
can
separate,
detect,
at
molecular
level.
However,
with
compromised
process.
The
concentration
ampholytes
required
mutually
exclusive
spectrometer
contamination.
We
have
improved
online
cIEF-ESI-MS
interface
reduce
(desalt)
amino
acid
in-line
after
In
proof
principle
experiments,
>90%
increase
area
under
curve
electropherograms
was
observed
compared
without
interface.
Protein
standards
including
cytochrome
C,
myoglobin,
α-casein
were
separated
resolved
high
reproducibility.
did
not
compromise
linearity
pH
gradient
separations,
achieving
R2
0.99.
addition,
tryptic
digest
BSA
demonstrates
baseline
resolution
as
little
0.02
pI
unit
difference
full
width
half-maximum
average
7.1
s.
Язык: Английский
Top-Down Proteomic Profiling of Protein Corona by High-Throughput Capillary Isoelectric Focusing-Mass Spectrometry
Reyhane Tabatabaeian Nimavard,
Seyed Amirhossein Sadeghi,
Morteza Mahmoudi
и другие.
Journal of the American Society for Mass Spectrometry,
Год журнала:
2025,
Номер
unknown
Опубликована: Март 2, 2025
In
the
rapidly
evolving
field
of
nanomedicine,
understanding
interactions
between
nanoparticles
(NPs)
and
biological
systems
is
crucial.
A
pivotal
aspect
these
formation
a
protein
corona
when
NPs
are
exposed
to
fluids
(e.g.,
human
plasma),
which
significantly
influences
their
behavior
functionality.
This
study
introduces
an
advanced
capillary
isoelectric
focusing
tandem
mass
spectrometry
(cIEF-MS/MS)
platform
designed
enable
high-throughput
reproducible
top-down
proteomic
analysis
corona.
Our
cIEF-MS/MS
technique
completed
each
within
30
min.
It
produced
proteoform
measurements
for
at
least
50
runs
regarding
proteoforms'
migration
time
[relative
standard
deviations
(RSDs)
<4%],
intensity
(Pearson's
correlation
coefficients
any
two
>0.90),
number
identifications
(71
±
10),
proteoform-spectrum
matches
(PrSMs)
(196
30).
Of
53
identified
genes,
33
potential
biomarkers
various
diseases
cancer,
cardiovascular
disease,
Alzheimer's
disease).
We
1-102
proteoforms
per
biomarker,
containing
sequence
variations
or
post-translational
modifications.
Delineating
in
by
our
fashion
will
benefit
nanobiointeractions
advance
both
diagnostic
therapeutic
nanomedicine
technologies.
Язык: Английский