Journal of Proteome Research,
Год журнала:
2023,
Номер
22(10), С. 3149 - 3158
Опубликована: Сен. 11, 2023
Mass
spectrometry
(MS)
enables
specific
and
accurate
quantification
of
proteins
with
ever-increasing
throughput
sensitivity.
Maximizing
this
potential
MS
requires
optimizing
data
acquisition
parameters
performing
efficient
quality
control
for
large
datasets.
To
facilitate
these
objectives
data-independent
(DIA),
we
developed
a
second
version
our
framework
data-driven
optimization
methods
(DO-MS).
The
DO-MS
app
v2.0
(do-ms.slavovlab.net)
allows
one
to
optimize
evaluate
results
from
both
label-free
multiplexed
DIA
(plexDIA)
supports
optimizations
particularly
relevant
single-cell
proteomics.
We
demonstrate
multiple
use
cases,
including
duty
cycle
methods,
peptide
separation,
number
survey
scans
per
cycle,
plexDIA
data.
interactive
display
generation
extensive
reports,
publication
figures
that
can
be
easily
shared.
source
code
is
available
at
github.com/SlavovLab/DO-MS.
Nature Communications,
Год журнала:
2023,
Номер
14(1)
Опубликована: Июль 12, 2023
Abstract
Liquid
chromatography
(LC)
coupled
with
data-independent
acquisition
(DIA)
mass
spectrometry
(MS)
has
been
increasingly
used
in
quantitative
proteomics
studies.
Here,
we
present
a
fast
and
sensitive
approach
for
direct
peptide
identification
from
DIA
data,
MSFragger-DIA,
which
leverages
the
unmatched
speed
of
fragment
ion
indexing-based
search
engine
MSFragger.
Different
most
existing
methods,
MSFragger-DIA
conducts
database
tandem
(MS/MS)
spectra
prior
to
spectral
feature
detection
peak
tracing
across
LC
dimension.
To
streamline
analysis
data
enable
easy
reproducibility,
integrate
into
FragPipe
computational
platform
seamless
support
library
building
DIA,
data-dependent
(DDA),
or
both
types
combined.
We
compare
other
tools,
such
as
DIA-Umpire
based
workflow
FragPipe,
Spectronaut,
DIA-NN
library-free,
MaxDIA.
demonstrate
fast,
sensitive,
accurate
performance
variety
sample
schemes,
including
single-cell
proteomics,
phosphoproteomics,
large-scale
tumor
proteome
profiling
Molecular & Cellular Proteomics,
Год журнала:
2023,
Номер
22(7), С. 100577 - 100577
Опубликована: Май 19, 2023
Accurate
biomarkers
are
a
crucial
and
necessary
precondition
for
precision
medicine,
yet
existing
ones
often
unspecific
new
have
been
very
slow
to
enter
the
clinic.
Mass
spectrometry
(MS)-based
proteomics
excels
by
its
untargeted
nature,
specificity
of
identification,
quantification,
making
it
an
ideal
technology
biomarker
discovery
routine
measurement.
It
has
unique
attributes
compared
affinity
binder
technologies,
such
as
OLINK
Proximity
Extension
Assay
SOMAscan.
In
in
previous
review
2017,
we
described
technological
conceptual
limitations
that
had
held
back
success.
We
proposed
'rectangular
strategy'
better
separate
true
minimizing
cohort-specific
effects.
Today,
this
converged
with
advances
MS-based
technology,
increased
sample
throughput,
depth
quantification.
As
result,
studies
become
more
successful,
producing
candidates
withstand
independent
verification
and,
some
cases,
already
outperform
state-of-the-art
clinical
assays.
summarize
developments
over
last
years,
including
benefits
large
cohorts,
which
acceptance.
Shorter
gradients,
scan
modes,
multiplexing
about
drastically
increase
cross-study
integration,
proxies
absolute
levels.
found
multiprotein
panels
inherently
robust
than
current
single
analyte
tests
capture
complexity
human
phenotypes.
Routine
MS
measurement
clinic
is
fast
becoming
viable
option.
The
full
set
proteins
body
fluid
(global
proteome)
most
important
reference
best
process
control.
Additionally,
increasingly
all
information
could
be
obtained
from
targeted
analysis
although
latter
may
straightforward
way
regular
use.
Many
challenges
remain,
not
least
regulatory
ethical
but
outlook
applications
never
brighter.
Nature Communications,
Год журнала:
2023,
Номер
14(1)
Опубликована: Сен. 22, 2023
Single-cell
resolution
analysis
of
complex
biological
tissues
is
fundamental
to
capture
cell-state
heterogeneity
and
distinct
cellular
signaling
patterns
that
remain
obscured
with
population-based
techniques.
The
limited
amount
material
encapsulated
in
a
single
cell
however,
raises
significant
technical
challenges
molecular
profiling.
Due
extensive
optimization
efforts,
single-cell
proteomics
by
Mass
Spectrometry
(scp-MS)
has
emerged
as
powerful
tool
facilitate
proteome
profiling
from
ultra-low
amounts
input,
although
further
development
needed
realize
its
full
potential.
To
this
end,
we
carry
out
comprehensive
orbitrap-based
data-independent
acquisition
(DIA)
for
proteomics.
Notably,
find
difference
between
optimal
DIA
methods
high-
low-load
samples.
We
improve
our
low-input
method
relying
on
high-resolution
MS1
quantification,
thus
enhancing
sensitivity
more
efficiently
utilizing
available
mass
analyzer
time.
With
input
tailored
method,
are
able
accommodate
long
injection
times
high
resolution,
while
keeping
the
scan
cycle
time
low
enough
ensure
robust
quantification.
Finally,
demonstrate
capability
approach
mouse
embryonic
stem
culture
conditions,
showcasing
global
proteomes
highlighting
differences
key
metabolic
enzyme
expression
subclusters.
Molecular & Cellular Proteomics,
Год журнала:
2024,
Номер
23(2), С. 100712 - 100712
Опубликована: Янв. 4, 2024
Data-independent
acquisition
(DIA)
mass
spectrometry
(MS)
has
emerged
as
a
powerful
technology
for
high-throughput,
accurate
and
reproducible
quantitative
proteomics.
This
review
provides
comprehensive
overview
of
recent
advances
in
both
the
experimental
computational
methods
DIA
proteomics,
from
data
schemes
to
analysis
strategies
software
tools.
are
categorized
based
on
design
precursor
isolation
windows,
highlighting
wide-window,
overlapping-window,
narrow-window,
scanning
quadrupole-based,
parallel
accumulation-serial
fragmentation
(PASEF)-enhanced
methods.
For
analysis,
major
classified
into
spectrum
reconstruction,
sequence-based
search,
library-based
de
novo
sequencing
sequencing-independent
approaches.
A
wide
array
tools
implementing
these
reviewed,
with
details
their
overall
workflows
scoring
approaches
at
different
steps.
The
generation
optimization
spectral
libraries,
which
critical
resources
also
discussed.
Publicly
available
benchmark
datasets
covering
global
proteomics
phosphoproteomics
summarized
facilitate
performance
evaluation
various
workflows.
Continued
synergistic
developments
versatile
components
expected
further
enhance
power
DIA-based
Nature Communications,
Год журнала:
2025,
Номер
16(1)
Опубликована: Янв. 2, 2025
Abstract
Data-independent
acquisition
has
become
a
widely
used
strategy
for
peptide
and
protein
quantification
in
liquid
chromatography-tandem
mass
spectrometry-based
proteomics
studies.
The
integration
of
ion
mobility
separation
into
data-independent
analysis,
such
as
the
diaPASEF
technology
available
on
Bruker’s
timsTOF
platform,
further
improves
accuracy
depth
achievable
using
acquisition.
We
introduce
diaTracer,
spectrum-centric
computational
tool
optimized
data.
diaTracer
performs
three-dimensional
(mass
to
charge
ratio,
retention
time,
mobility)
peak
tracing
feature
detection
generate
precursor-resolved
“pseudo-tandem
spectra”,
facilitating
direct
(“spectral-library
free”)
identification
from
is
stand-alone
fully
integrated
FragPipe
platform.
demonstrate
performance
data
triple-negative
breast
cancer,
cerebrospinal
fluid,
plasma
samples,
phosphoproteomics
human
leukocyte
antigens
immunopeptidomics
experiments,
low-input
spatial
study.
also
show
that
enables
unrestricted
post-translational
modifications
open/mass-offset
searches.
The
ability
to
map
a
proteomic
fingerprint
transcriptomic
data
would
master
the
understanding
of
how
gene
expression
translates
into
actual
phenotype.
In
contrast
nucleic
acid
sequencing,
in
vitro
protein
amplification
is
impossible
and
no
single
cell
workflow
has
been
established
as
gold
standard
yet.
Advances
microfluidic
sample
preparation,
multi-dimensional
separation,
sophisticated
acquisition
strategies,
intelligent
analysis
algorithms
have
resulted
major
improvements
successfully
analyze
such
tiny
amounts
with
steadily
boosted
performance.
However,
among
broad
variation
published
approaches,
it
commonly
accepted
that
highest
possible
sensitivity,
robustness,
throughput
are
still
most
urgent
needs
for
field.
While
many
labs
focused
on
multiplexing
achieve
these
goals,
label-free
SCP
highly
promising
strategy
well
whenever
high
dynamic
range
unbiased
accurate
quantification
needed.
We
here
focus
recent
advances
single-cell
mass
spectrometry
workflows
try
guide
our
readers
choose
best
method
or
combinations
methods
their
specific
applications.
further
highlight
which
techniques
propitious
future
applications
but
also
limitations
we
foresee
bioRxiv (Cold Spring Harbor Laboratory),
Год журнала:
2023,
Номер
unknown
Опубликована: Фев. 2, 2023
Abstract
Data-independent
acquisition
(DIA)
approaches
provide
comprehensive
records
of
all
detectable
pre-cursor
and
fragment
ions.
Here
we
introduce
midiaPASEF,
a
novel
DIA
scan
mode
using
mobility-specific
micro-encoding
overlapping
quadrupole
windows
to
optimally
cover
the
ion
population
in
mobility-mass
charge
plane.
Using
mobility-encoded
windows,
midiaPASEF
maximizes
information
content
acquisitions
which
enables
determination
precursor
m/z
each
with
precision
less
than
2
Th.
The
Snakemake-based
MIDIAID
pipeline
integrates
algorithms
for
multidimensional
peak
detection
machine-learning-based
classification
precursor-fragment
relationships.
fully
automated
processing
deconvolution
midia-PASEF
files
exports
highly
specific
DDA-like
MSMS
spectra
are
suitable
de
novo
sequencing
can
be
searched
directly
established
tools
including
PEAKS,
FragPipe
Mascot.
identifies
over
40
unique
peptides
per
second
provides
powerful
library-free
analyses
phosphopeptidome
immunopeptidome
samples.
