3.6 In Vivo Mouse
Imaging
For each experiment, imaging sessions should be scheduled as
appropriate. The first one is realized 30 min post-infection (p.i.)
to ensure that the inoculum has been correctly injected. During the
acute phase of infection (from day 1 to day 5 p.i.), imaging sessions
are usually performed every day in ventral position to follow disease
progression in vivo (see Note 6). From day 14 p.i., the level of renal
colonization can be monitored in the dorsal position, with imaging
sessions at determined time points. For any experiment, an image
should be acquired with control mice to establish luminescence
background.
1. Thaw aliquots of D-luciferin and warm to room temperature,
as many aliquots as experimental groups. Luciferin injection
and anesthesia are performed sequentially.
2. Ten to twelve minutes before imaging, inject 100 μL of Dluciferin in the peritoneal cavity of the first experimental
group of mice to be imaged.
3. Place mice in the induction chamber and maintain them under
anesthesia for 10 min (see Note 7).
4. Transfer each anesthetized mouse (be careful to handle infected
mice on a tissue to prevent the urine from contaminating the
environment) immediately to the dark box imaging system on a
disposable black sheet (to avoid cross contamination), and
position the head gently in the transparent cones inserted in
the anesthesia manifold to maintain the anesthesia. Depending
on the type of imaging, place animals in ventral or dorsal
position (see Note 8). Center the animals between each divider
and align the legs and tail of each mouse. Pay attention that the
order of individual mice should be respected for individual
follow-up over time.
5. Close the door of the dark box and click Acquire on the control
screen. First, acquire in automatic exposure time to get the
absolute light value and then with a 5 min exposure time for
a picture (see Note 9). When the first image capture is activated,
autosave function is proposed to the user. Select Yes to save all
images in a folder specifically created for the experiment.
6. While the first group of mice is imaged, inject the luciferin to
the second group of mice, and put them under anesthesia for
10 min before imaging.
7. After imaging, remove mice from the dark box system, and
return them to their respective cage, in the security position, on
their right side. Monitor for complete recovery from anesthesia
(approximately 2 min for healthy mice) before putting the cage
back on the ventilated rack.
In Vivo Imaging of Bioluminescent Leptospires
155
Imaging
For each experiment, imaging sessions should be scheduled as
appropriate. The first one is realized 30 min post-infection (p.i.)
to ensure that the inoculum has been correctly injected. During the
acute phase of infection (from day 1 to day 5 p.i.), imaging sessions
are usually performed every day in ventral position to follow disease
progression in vivo (see Note 6). From day 14 p.i., the level of renal
colonization can be monitored in the dorsal position, with imaging
sessions at determined time points. For any experiment, an image
should be acquired with control mice to establish luminescence
background.
1. Thaw aliquots of D-luciferin and warm to room temperature,
as many aliquots as experimental groups. Luciferin injection
and anesthesia are performed sequentially.
2. Ten to twelve minutes before imaging, inject 100 μL of Dluciferin in the peritoneal cavity of the first experimental
group of mice to be imaged.
3. Place mice in the induction chamber and maintain them under
anesthesia for 10 min (see Note 7).
4. Transfer each anesthetized mouse (be careful to handle infected
mice on a tissue to prevent the urine from contaminating the
environment) immediately to the dark box imaging system on a
disposable black sheet (to avoid cross contamination), and
position the head gently in the transparent cones inserted in
the anesthesia manifold to maintain the anesthesia. Depending
on the type of imaging, place animals in ventral or dorsal
position (see Note 8). Center the animals between each divider
and align the legs and tail of each mouse. Pay attention that the
order of individual mice should be respected for individual
follow-up over time.
5. Close the door of the dark box and click Acquire on the control
screen. First, acquire in automatic exposure time to get the
absolute light value and then with a 5 min exposure time for
a picture (see Note 9). When the first image capture is activated,
autosave function is proposed to the user. Select Yes to save all
images in a folder specifically created for the experiment.
6. While the first group of mice is imaged, inject the luciferin to
the second group of mice, and put them under anesthesia for
10 min before imaging.
7. After imaging, remove mice from the dark box system, and
return them to their respective cage, in the security position, on
their right side. Monitor for complete recovery from anesthesia
(approximately 2 min for healthy mice) before putting the cage
back on the ventilated rack.
In Vivo Imaging of Bioluminescent Leptospires
155