Seburn2_Protocol
Project
protocol
—
Contents
Workflow
and
sampling
Equipment
Reagents,
supplies,
and
solutions
Procedure
for
gait
analysis
Procedure
for
grip
strength
measurement
Data
Definitions
References
Workflow
and
sampling
Workflow
Step |
Procedure
performed |
Equipment |
Data
collected |
|
Capture
images
of
mice
walking |
mouse
treadmill
system;
digital
video
recorder |
- |
2 |
Gait
analysis
is
conducted
by
converting
raw
digital
images
into
MPEG
format
that
can
be
exported
into
Treadscan™
software |
computer
|
stance,
swing,
and
stride
times;
stride
length
and
dynamic
stance
(track)
width |
* |
Grip
strength
is
measured |
grip
strength
meter |
peak
tension |
*
Grip
strength
measurements
were
conducted
separately
before
or
after
gait
analysis.
Equipment
•
Gait
analysis
system-
hardware
(Prototype)
-
The
system
is
configured
to
allow
video
capture
from
below
the
mice
during
treadmill
locomotion.
Hardware
consists
of
a
treadmill
with
a
clear
transparent
belt
and
a
digital
video
camera
(Basler
A301fc)
mounted
below.
A
rectangular,
opaque
plexiglas
testing
chamber
(20
x
4
x
16.5
cm)
is
mounted
over
the
tread
and
serves
to
keep
the
mouse
within
view
of
the
camera.
Illumination
of
the
ventral
side
of
the
mouse
is
adjusted
to
obtain
optimal
contrast
between
the
paws
and
body
of
the
mouse.
The
digital
camera
operates
at
100
frames
per
seconds
(fps)
with
a
resolution
of
658
x
494
pixels
and
the
camera
output
is
fed
directly
(Firewire
IEEE-1394)
to
a
2.4GHz,
1GB
RAM
computer.
-
Digital
video
recording
software
is
used
to
capture
the
raw
video
images
and
to
convert
them
to
MPEG
format.
The
MPEG
file
is
then
exported
to
custom
analysis
software
(Treadscan™
1.0,
Clever
Sys,
Inc.,
Reston,
VA,
USA)
•
Gait
analysis
software
system
(CleverSys
Inc.,
USA)
The
Treadscan™
software
determines
when
individual
paws
are
in
contact
with
the
treadmill
and
then
uses
this
determination
to
derive
more
than
20
different
measures
including
standard
time-domain
gait
parameters
(stride,
stance
and
swing
time). Analysis
is
interactive
and
provides
frame-by-frame
feedback
that
allows
the
user
to
confirm
that
the
mouse
is
walking
with
a
consistent
pattern
and
that
the
paws
are
tracked
accurately.
Measurements
of
improper
strides
(e.g.
paw
not
being
tracked,
mouse
moving
forward
or
back
on
tread)
can
be
flagged
and
excluded
from
the
data
output.
•
Grip
strength
meter,
Columbus
Instruments
Reagents,
supplies,
solutions
Acclimation
to
test
conditions
In
general
all
mice
are
acclimated
for
at
least
1
h
in
the
testing
room.
Procedure
for
gait
analysis
a.
Standard
video
capture
protocol.
Camera
settings,
lighting
and
treadmill
speed
(20
cm/s
for
all
studies)
are
preset
and
background
images
including
distance
calibration
are
captured,
prior
to
introduction
of
the
mouse.
b.
Each
mouse
is
placed
into
the
testing
chamber
where
it
could
be
seen
live
on
the
computer
screen
and
within
15
s
the
treadmill
is
turned
on
and
operating
at
20
cm/s.
c.
The
mouse
walking
is
then
recorded
for
a
preset
fixed
number
of
frames
(796
frames)
after
which
the
treadmill
is
stopped
and
the
mouse
removed
from
the
testing
chamber.
d.
Each
mouse
is
in
the
testing
chamber
for
a
maximum
total
of
45-60
s.
e.
A
step
cycle
or
stride
consists
of
two
consecutive
contacts
of
a
given
foot.
At
least
5
strides
for
each
foot
is
included
for
a
given
data
set.
Only
strides
recorded
when
the
mice
are
walking
in
a
fixed
position
relative
(i.e.
at
the
preset
speed)
to
the
camera
are
included.
Whenever
possible
consecutive
strides
are
used.
However,
in
the
presence
of
intervening
aberrant
strides
(e.g.
jumps
or
partial
steps),
as
do
periods
of
"stop
and
go"
walking,
data
from
most
mice
included
2
or
3
sets
of
consecutive
strides
for
a
given
trial.
f.
Stored
video
recordings
are
subsequently
analyzed
to
derive
standard
gait
indices
including
stride,
stance,
and
swing
time.
g.The
tread
is
cleaned
between
trials.
Investigator's
note:
The
gait
analysis
device
has
been
fully
validated
and
the
results
have
been
published
(Wooley
et.
al.,
2005).
Procedure
for
measuring
forelimb
grip
strength
a.
Each
mouse
is
gently
lowered
over
the
grip
strength
meter
forelimb
pull
bar
assembly
(76
mm
x
50
mm),
such
that
only
the
front
paws
are
able
to
grip
the
trapeze.
b.
The
speed
and
angle
of
pull
manually
applied
to
the
mouse
are
standardized
to
reduce
variability
between
measurements.
Therefore,
the
body
of
the
mouse
is
kept
at
a
vertical
angle,
with
a
given
standard
weight
applied
(or
taped
to
the
tail)
in
a
vertical
direction
to
"break"
the
mouse's
grip
and
to
produce
the
peak
force
or
tension.
c.
Three
consecutive
forelimb
grip
strength
trials
are
performed
and
recorded.
d.
Any
morphological
defect
or
signs
of
inability
to
grip
and
successfully
perform
the
test
is
noted.
e.
The
Trapeze
is
cleaned
after
each
mouse
tested.
Data
collected
by
investigator
Grip
strength
(peak
tension)
in
6,
12,
18,
and
24
mo
old
mice.
Gait
analysis
of
the
front
and
rear
paws
(speed,
distance,
and
frequency
of
stride)
in
6
and
18
mo
old
mice.
|
|
Test age:
6, 12, 18, 24 mo
