Molecular & Cellular Proteomics,
Journal Year:
2024,
Volume and Issue:
unknown, P. 100880 - 100880
Published: Nov. 1, 2024
Mass
spectrometry-based
proteome
profiling
of
trace
analytes
including
single
cells
benefits
from
liquid
chromatography
separations
operated
at
low
flow
rates
(e.g.,
<50
nL/min).
However,
high-pressure
binary
pumps
needed
to
achieve
such
are
not
commercially
available,
and
instead
require
splitting
the
gradient
low-nanoliter-per-minute
rates.
Gradient
can
waste
solvent
lead
inconsistencies.
To
address
this,
we
have
developed
a
method
for
creating
gradients
by
combining
plugs
mobile
phase
increasing
strength
together
in
column,
then
relying
on
Taylor
dispersion
form
desired
smooth
profile.
Additionally,
our
dramatically
reduces
costs,
as
only
isocratic
pump
is
required.
Following
development
profiles
both
10-
20-min
active
gradients,
measured
200
pg
injections
HeLa
digest
using
timsTOF
mass
spectrometer.
Finally,
investigated
differences
protein
expression
between
originating
two
different
colonies
ATG-knockout
cells.
Thousands
proteins
were
quantified,
potential
mechanism
explaining
differential
immune
responses
these
upon
exposure
viral
DNA
treatment
was
determined.
Nature Communications,
Journal Year:
2024,
Volume and Issue:
15(1)
Published: March 20, 2024
Mass
spectrometry
(MS)-based
proteomics
workflows
typically
involve
complex,
multi-step
processes,
presenting
challenges
with
sample
losses,
reproducibility,
requiring
substantial
time
and
financial
investments,
specialized
skills.
Here
we
introduce
One-Tip,
a
methodology
that
seamlessly
integrates
efficient,
one-pot
preparation
precise,
narrow-window
data-independent
acquisition
(nDIA)
analysis.
One-Tip
substantially
simplifies
processing,
enabling
the
reproducible
identification
of
>9000
proteins
from
~1000
HeLa
cells.
The
versatility
is
highlighted
by
nDIA
~6000
in
single
cells
early
mouse
embryos.
Additionally,
study
incorporates
Uno
Single
Cell
Dispenser™,
demonstrating
capability
single-cell
>3000
identified
per
cell.
We
also
extend
workflow
to
analysis
extracellular
vesicles
(EVs)
extracted
blood
plasma,
its
high
sensitivity
identifying
16
ng
EV
preparation.
expands
capabilities
proteomics,
offering
greater
depth
throughput
across
range
types.
Nature Methods,
Journal Year:
2025,
Volume and Issue:
unknown
Published: Jan. 16, 2025
Single-cell
proteomics
(SCP)
promises
to
revolutionize
biomedicine
by
providing
an
unparalleled
view
of
the
proteome
in
individual
cells.
Here,
we
present
a
high-sensitivity
SCP
workflow
named
Chip-Tip,
identifying
>5,000
proteins
HeLa
It
also
facilitated
direct
detection
post-translational
modifications
single
cells,
making
need
for
specific
modification-enrichment
unnecessary.
Our
study
demonstrates
feasibility
processing
up
120
label-free
samples
per
day.
An
optimized
tissue
dissociation
buffer
enabled
effective
single-cell
disaggregation
drug-treated
cancer
cell
spheroids,
refining
overall
analysis.
Analyzing
nondirected
human-induced
pluripotent
stem
differentiation,
consistently
quantified
markers
OCT4
and
SOX2
cells
lineage
such
as
GATA4
(endoderm),
HAND1
(mesoderm)
MAP2
(ectoderm)
different
embryoid
body
sets
benchmark
sensitivity
throughput,
with
broad
applications
basic
biology
identification
type-specific
therapeutic
targets.
Communications Biology,
Journal Year:
2025,
Volume and Issue:
8(1)
Published: Jan. 17, 2025
Recent
developments
in
mass
spectrometry-based
proteomics
have
established
it
as
a
robust
tool
for
system-wide
analyses
essential
pathophysiological
research.
While
post-mortem
samples
are
critical
source
these
studies,
our
understanding
of
how
body
decomposition
influences
the
proteome
remains
limited.
Here,
we
revisited
published
data
and
conducted
clinically
relevant
time-course
experiment
mice,
revealing
organ-specific
regulation
after
death,
with
only
fraction
changes
linked
to
protein
autolysis.
The
liver
spleen
exhibit
significant
proteomic
alterations
within
hours
post-mortem,
whereas
heart
displays
modest
changes.
Additionally,
subcellular
compartmentalization
leads
an
unexpected
surge
at
earliest
interval
(PMI).
comprehensive
analysis
semi-tryptic
peptides,
distinct
consensus
motifs
different
organs,
indicating
protease
activity.
In
conclusion,
findings
emphasize
importance
considering
PMI
effects
when
designing
may
significantly
overshadow
impacts
diseases.
Preferably,
should
be
taken
operation
room,
especially
studies
including
or
trans-organ
comparison.
single-organ
planning
involve
careful
control
PMI.
Molecular & Cellular Proteomics,
Journal Year:
2024,
Volume and Issue:
23(5), P. 100750 - 100750
Published: March 20, 2024
Spatial
tissue
proteomics
integrating
whole-slide
imaging,
laser
microdissection,
and
ultrasensitive
mass
spectrometry
is
a
powerful
approach
to
link
cellular
phenotypes
functional
proteome
states
in
(patho)physiology.
To
be
applicable
large
patient
cohorts
low
sample
input
amounts,
including
single-cell
applications,
loss-minimized
streamlined
end-to-end
workflows
are
key.
We
here
introduce
an
automated
preparation
protocol
for
microdissected
samples
utilizing
the
cellenONE
robotic
system,
which
has
capacity
process
192
3
h.
Following
microdissection
collection
directly
into
proteoCHIP
LF
48
or
EVO
96
chip,
our
optimized
facilitates
lysis,
formalin
de-crosslinking,
tryptic
digest
of
low-input
archival
samples.
The
seamless
integration
with
Evosep
ONE
LC
system
by
centrifugation
allows
'on-the-fly'
clean-up,
particularly
pertinent
workflows.
validate
method
human
tonsil
tissue,
where
we
profile
proteomes
spatially-defined
B-cell,
T-cell,
epithelial
microregions
4000
μm
Analytical Chemistry,
Journal Year:
2025,
Volume and Issue:
unknown
Published: Jan. 6, 2025
Single-cell
proteomics
(SCP)
detected
based
on
different
technologies
always
involves
batch-specific
variations
because
of
differences
in
sample
processing
and
other
potential
biases.
How
to
integrate
SCP
data
effectively
has
become
a
great
challenge.
Integration
not
only
requires
the
conservation
true
biological
variances,
but
also
realizes
removal
unwanted
batch
effects.
In
this
study,
benchmarking
analysis
popular
integration
methods
was
conducted
determine
most
suitable
method
for
data.
To
comprehensively
evaluate
performance
these
methods,
novel
evaluation
system
proposed
integrating
This
consists
three
objective
measures
from
perspectives:
category
(a),
efficacy
correcting
effects;
(b),
power
conserving
variances;
(c),
ability
identify
consistent
markers.
For
comprehensive
evaluation,
five
benchmark
sets
under
scenarios
(containing
substantial
proteins,
cells,
multiple
batches,
cell
types,
unbalanced
data)
were
utilized
selecting
method.
As
result,
ComBat,
Scanorama,
Seurat
version
3
CCA,
identified
as
recommended
Overall,
systematic
might
provide
valuable
guidance
choosing
appropriate
SCP.
Journal of Proteome Research,
Journal Year:
2025,
Volume and Issue:
unknown
Published: Feb. 3, 2025
Mass
spectrometry-based
single-cell
proteomics
(SCP)
is
gaining
momentum
but
remains
limited
to
a
few
laboratories
due
the
high
costs
and
specialized
expertise
required.
The
ability
send
samples
core
facilities
would
benefit
nonspecialist
popularize
SCP
for
biological
applications.
However,
no
methods
have
been
tested
in
"freeze"
proteome
state
while
maintaining
cell
integrity
transfer
between
or
prolonged
sorting
using
fluorescence-activated
(FACS).
This
study
evaluates
whether
short-term
formaldehyde
(FA)
fixation
can
maintain
states.
We
demonstrate
that
FA
does
not
substantially
affect
protein
recovery,
even
without
heating
strong
detergents,
maintains
analytical
depth
compared
with
classical
workflows.
Fixation
also
preserves
drug-induced
specific
perturbations
of
abundance
during
sample
preparation
analysis.
Our
findings
suggest
facilitate
by
enabling
shipping
sorting,
potentially
democratizing
access
technology
expanding
its
application
research,
thereby
accelerating
discoveries
biology
personalized
medicine.
Journal of Integrative Plant Biology,
Journal Year:
2025,
Volume and Issue:
unknown
Published: March 10, 2025
Small
peptides
(SPs)
are
pivotal
signaling
molecules
that
play
essential
roles
in
the
precise
regulation
of
plant
growth,
development,
and
stress
responses.
Recent
advancements
sequencing
technologies,
bioinformatics
approaches,
biochemical
molecular
techniques
have
significantly
enhanced
accuracy
SP
identification,
unveiling
their
diverse
biological
functions
plants.
This
review
provides
a
comprehensive
overview
characteristics
methodologies
for
identifying
SPs
It
highlights
recent
discoveries
regarding
pathways
regulating
plant-microbial
interactions,
as
well
contributions
to
resilience
under
various
environmental
stresses,
including
abiotic
stress,
nutrient
deficiencies,
biotic
challenges.
Additionally,
we
discuss
current
insights
into
potential
applications
outline
future
research
directions
aimed
at
leveraging
these
enhance
adaptation
By
integrating
findings,
this
lays
foundation
advancing
understanding
utilization
improve
productivity.
