Project
protocol
—
Contents
Workflow
and
sampling
Equipment
Reagents,
supplies,
and
solutions
Procedure
1:
Conducting
behavioral
tests
Procedure
2:
Corticosterone
radioimmunoassay
Data
References
Workflow
and
sampling
Workflow
|
Procedure
performed |
Equipment |
Duration
(min) |
Age
(wk) |
Data
collected |
1a |
Mice
are
assessed
for
the
tail
suspension
test |
Tail
suspension
apparatus |
10 |
5-18 |
|
1b |
Body
weights
are
obtained
from
each
mouse
after
tail
suspension
test |
balance
scale |
- |
|
|
2 |
Exploratory
behavior
is
assessed
in
an
open
field
arena |
Open
field
arena |
7 |
|
percent
center
occupancy,
total
activity
|
3 |
Blood
is
collected
for
serum
corticosterone
|
radioimmunoassay
kit |
- |
5-18 |
|
Equipment
•
Mouse
Tail
Suspension
setup
(Med
Associates,
Georgia,
VT)
see
Figure
1
below
•
Open
field
arena:
Med
Associates
mouse
Open
Field
contained
within
an
environmental
chamber
that
provides
white
noise
and
low,
indirect
lighting.
This
package
includes
a
10.75"
x
10.75"
(27.3
cm
x
27.3
cm)
test
environment
with
three
16
beam
infrared
(I/R)
arrays,
48
channel
control,
cables,
interface
card,
power
supply,
and
single
station
software.
Up
to
eight
test
stations
may
be
attached
to
a
single
computer.
These
boxes
measure
activity
using
infrared
beam
breaks
on
the
x,
y,
and
z
axes
(see
Figure
2
below).
•
Computer
•
Freezer
at
-80°C
•
Dissecting
kit:
decapitator
Figure
1.
Schematic
layout
of
the
tail
suspension
test
(not
drawn
to
scale).
Figure
2.
Schematic
layout
of
the
open
field
arena.
Reagents,
supplies,
solutions
•
Double
antibody
RIA
kit
for
serum
corticosterone
(Diagnostics
Products
Corp.
Los
Angeles,
CA)
•
Ice
and
ice
bucket
•
Disinfectant
solution
•
Paper
towels
and
other
cleaning
materials
•
Blood
collection
tubes
Procedure
1:
Conducting
behavioral
tests
Common
procedures
a.
Mice
are
brought
from
separate
colony
rooms
to
the
testing
room
before
behavioral
survey
is
done.
b.
Behavioral
testing
is
conducted
between
1300h
and
1600h.
c.
Strains
are
randomized
across
day,
time
of
testing,
and
equipment.
d.
Four
to
eight
mice
are
tested
at
a
time,
and
each
individual
is
only
tested
once
in
each
task.
e.
The
behavioral
apparatus
is
cleaned
with
a
disinfectant
between
each
test,
and
at
the
end
of
each
survey,
mice
are
returned
to
their
home
cages.
Tail
suspension
test
a.
Tail
Suspension
Test
(TST)
is
performed
using
a
Med
Associates
Mouse
setup.
b.
In
this
test,
the
mouse
is
placed
inside
a
3-sided
cubicle
and
suspended
by
its
tail
from
a
hanger
attached
to
a
precision
linear
load
cell
that
measures
activity.
c.
Mice
are
acclimated
to
testing
conditions
for
at
least
1
min,
this
is
to
account
for
the
fact
that
mice
are
uniformly
active
during
this
time.
d.
Measurements
are
taken
in
200-ms
increments
for
7
min,
with
threshold
=
3
and
gain
=
8.
e.
Percent
immobility
is
calculated
by
determining
the
time
spent
immobile
during
the
last
6
min
of
the
test,
and
reported
as
percent
of
time
(on
a
scale
of
0-1)
spent
immobile
during
the
TST.
f.
Mice
that
climbed
their
tail
or
fell
off
the
hanger
are
excluded
from
analysis.
g.
Body
weights
are
obtained
immediately
after
the
TST,
at
approximately
7-10
wks
of
age.
Open
field
test
a.
One
week
after
tail
suspension
test,
mice
are
tested
in
a
Med
Associates
mouse
Open
Field
Test
(OFT)
contained
within
an
environmental
chamber
that
provides
white
noise
and
low,
indirect
lighting.
b.
Mice
are
placed
in
the
center
of
the
field
and
allowed
to
explore
freely
for
10
min.
c.
Center
time
is
calculated
as
the
percent
of
time
(on
a
scale
of
0-1)
spent
in
the
center
25%
of
the
open
field
arena.
d.
Activity
is
measured
by
total
distance
traveled
during
the
10
min
OFT.
Procedure
2:
Corticosterone
Radioimmunoassay
a.
All
collections
are
performed
between
1100h
and
1500h.
b.
Trunk
blood
is
collected
within
30
s
of
removing
the
mice
from
the
colony
room
and
following
immediate
decapitation.
c.
Blood
is
allowed
to
clot
on
ice,
and
then
serum
is
separated
out
and
stored
at
-80°C.
d.
Corticosterone
is
measured
from
10
mL
of
serum
using
a
double
antibody
RIA
kit.
e.
The
assay
is
calibrated
to
within
a
minimum
detectable
level
of
0.5
mg/dL
of
serum
corticosterone,
and
interassay/intrassay
coefficients
of
variance
of
12.48%
and
6.11%,
respectively.
f.
The
RIA
is
performed
by
the
Ligand
Assay
Core
at
Northwestern
University.
Data
collected
and
submitted
by
investigator
Open
field
test:
percent
center
occupancy
and
total
activity.
Tail
suspension
test:
percent
immobility.
Serum
corticosterone
only
Test age:
7-10wks*
