Analytical Methods,
Journal Year:
2024,
Volume and Issue:
16(20), P. 3220 - 3230
Published: Jan. 1, 2024
Tuberculosis
caused
by
Mycobacterium
bovis
poses
a
global
infectious
threat
to
humans
and
animals.
Therefore,
there
is
an
urgent
need
develop
sensitive,
precise,
easy-to-readout
strategy.
Here,
novel
tandem
combination
of
CRISPR/Cas12a
system
with
dual
HCR
(denoted
as
CRISPR/Cas12a-D-HCR)
was
constructed
for
detecting
bovis.
Based
on
the
efficient
trans-cleavage
activity
active
system,
tandem-dsDNA
PAM
sites
established
using
two
flexible
hairpins,
providing
multiple
binding
further
amplification.
Furthermore,
activation
Cas12a
initiated
second
hybridization
chain
reaction
(HCR),
which
integrated
complete
G-quadruplex
sequences
assemble
hemin/G-quadruplex
DNAzyme.
With
addition
H2O2
ABTS,
colorimetric
signal
readout
strategy
achieved.
Consequently,
CRISPR/Cas12a-D-HCR
achieved
satisfactory
detection
linear
range
from
20
aM
50
fM,
limit
low
2.75
single
mismatched
recognition
capability,
demonstrating
good
discrimination
different
bacterial
species.
Notably,
practical
application
performance
verified
via
standard
method,
recovery
ranging
96.0%
105.2%
relative
deviations
(RSD)
0.95%
6.45%.
The
proposed
sensing
served
promising
accurate
in
food
safety
agricultural
fields.
Analytical Chemistry,
Journal Year:
2024,
Volume and Issue:
unknown
Published: Sept. 25, 2024
Current
nucleic
acid-responsive
DNA
hydrogels
face
significant
challenges,
such
as
the
requirement
for
high
target
concentrations,
frequent
redesigns,
and
increased
costs,
which
limit
their
practical
applications
in
biosensing.
To
address
these
issues,
we
developed
a
novel
biosensing
platform
integrating
CRISPR/Cas12a
system
into
an
RCA-based
hydrogel.
The
hydrogel
used
could
preencapsulate
diverse
signal
molecules
comprising
GelRed,
methylene
blue,
gold
nanoparticles,
were
released
upon
Cas12a-mediated
cleavage.
This
design
enabled
customizable
output,
including
fluorescence,
electrochemistry,
colorimetry,
thereby
ensuring
platform's
adaptability
to
various
detection
scenarios.
Our
was
highly
specific
methicillin-resistant
Journal of Agricultural and Food Chemistry,
Journal Year:
2023,
Volume and Issue:
72(1), P. 874 - 882
Published: Dec. 29, 2023
The
sensitive
and
accurate
detection
of
ochratoxin
A
(OTA)
is
crucial
for
public
health
due
to
its
high
toxicity.
Herein,
using
Au
nanoparticle
(NP)-attached
CdS/UiO-66-NH2
heterostructures
as
photoactive
materials,
a
photoelectrochemical
(PEC)
aptasensor
was
presented
the
ultrasensitive
assay
OTA
based
on
competitive
displacement
reaction
triggering
trans-cleavage
ability
CRISPR/Cas12a.
In
this
sensing
strategy,
methylene
blue-labeled
single-stranded
DNA
(MB-ssDNA)
immobilized
NPs/CdS/UiO-66-NH2
electrode
accelerate
separation
photogenerated
carrier,
thus
producing
significantly
increased
PEC
response.
presence
OTA,
it
specifically
bound
with
aptamer
(Apt)
resulted
in
release
activation
chain,
characteristics
MB-ssDNA
cut
randomly
surface
convert
signal
from
"on"
"off"
state,
thereby
achieving
quantitative
OTA.
CRISPR/Cas12a-derived
exhibited
excellent
sensitivity
specificity,
linear
range
100
50
ng/mL
limit
38
fg/mL.
Overall,
proposed
could
provide
rapid,
accurate,
method
determination
actual
samples.
Journal of Nanobiotechnology,
Journal Year:
2023,
Volume and Issue:
21(1)
Published: Dec. 20, 2023
A
multimodal
analytical
strategy
utilizing
different
modalities
to
cross-validate
each
other,
can
effectively
minimize
false
positives
or
negatives
and
ensure
the
accuracy
of
detection
results.
Herein,
we
establish
a
colorimetric,
photothermal,
fluorescent
triple
modal
CRISPR/Cas12a
platform
(CPF-CRISPR).
An
MNPs-ssDNA-HRP
signal
probe
is
designed
act
as
substrate
trigger
three
outputs.
In
presence
DNA
target,
cleaved
by
activated
CRISPR/Cas12a,
resulting
in
release
HRP
generating
short
strands
with
3-terminal
hydroxyl
on
magnetic
beads.
The
released
subsequently
catalyzed
TMB-H2O2
reaction
oxidized
TMB
used
for
colorimetric
photothermal
detection.
Under
catalysis
terminal
deoxynucleotidyl
transferase
(TdT),
remaining
are
primers
form
poly-T
function
scaffolds
copper
nanoclusters
output.
To
verify
practical
application
CPF-CRISPR,
employed
MRSA
model.
results
demonstrate
platform's
high
sensitivity,
limit
101
CFU/mL
when
combined
recombinase
polymerase
amplification.
Therefore,
harnessing
programmability
biosensor
has
potential
detect
various
drug-resistant
bacteria,
demonstrating
significant
applicability.
Achieving
effective
control
over
microbial
contamination
necessitates
the
precise
and
concurrent
identification
of
numerous
pathogens.
In
this
research,
we
have
devised
a
remarkably
sensitive
duplex
droplet
digital
PCR
(dddPCR)
reaction
system
to
simultaneously
detect
Pseudomonas
aeruginosa
(P.
aeruginosa)
fragi
fragi).
Employing
comparative
genomics,
identified
four
genes
P.
fragi.
By
specific
analysis,
RS22680
gene
was
selected
as
detection
target
LasR
chosed
aeruginosa,
which
were
applied
construct
dddPCR
reaction.
terms
specificity,
sensitivity
anti-interference
ability,
constructed
verified
analyzed.
The
assay
showed
excellent
applicability,
evidenced
by
limit
100
CFU/mL.
When
concentration
natural
background
bacteria
in
milk
or
fresh
meat
times
that
bacteria,
method
still
capable
completing
absolute
quantification.
simulation
actual
sample
contamination,
could
be
detected
after
3
h
enrichment
culture,
6
h.
established
ddPCR
exhibits
exceptional
performance,
serving
foundation
for
simultaneous
various
pathogenic
food
products.
