35. Visually inspect the silver block to ensure that it is clean and
dry. Dust particles can be removed using a bulb air blower.
36. Replace the cryomicroscope stage lid to minimize dust ingress.
It is not necessary to tighten the lid completely.
37. If additional experiments are planned, reload the previously
saved temperature profile (Table 1) into the Linksys software
(see Note 97). When saving of the current video has completed
(see Note 95), repeat the above protocols starting with the
sample preparation procedure (Subheading 3.5).
3.8 Experiment
Shutdown
1. Set the stage temperature to a value in the range 30–40
C, by
modifying the active row of the temperature profile table in
Linksys (see Note 39); specify an indefinite hold (e.g., set
Time ¼ 99,999).
2. Maximize clearance between the BCS196 lid and the microscope objectives by using the coarse focus controls to lower the
stage (see Notes 49 and 50).
3. Detach the LNP purge tubing extension from the BCS196
front gas port by pushing the valve sleeve toward the stage
body until the valve clicks shut.
4. Disconnect the LNP liquid nitrogen withdrawal tubing from
the coolant exhaust pipe on the BCS196 stage (see Note 98).
5. Disconnect the dewar’s insulated capillary tubing from the
coolant inlet pipe on the BCS196 stage (see Notes 98 and 99).
6. Terminate control of the stage temperature, by clicking the
blue square button in the Temperature Control toolbar in the
Linksys interface to stop execution of the temperature profile.
7. Select “Disconnect” from the “File” menu; confirm that the
temperature display reads “No Comm.”
8. Exit the Linksys32 software, and then power down the liquid
nitrogen pump and the cryomicroscope system controller.
9. Unclasp the latches of the lid on the LNP dewar, and gently
remove the lid. If necessary, discard any leftover liquid nitrogen
using appropriate disposal methods.
10. Detach the LNP window tubing from the BCS196 stage lid,
and remove the metal clip from the end of the tubing.
11. Disconnect the LEMO connector from the socket on the left
side of the BCS196 stage body, by pulling it straight out.
12. When all image and videos from the experiments have been
saved, close the camera control software and shut down the
computer.
13. Power down the camera by disconnecting its power adapter.
14. Disconnect the data transfer cable and the trigger interface
cable from the camera.
High-Speed Video Cryomicroscopy
239
dry. Dust particles can be removed using a bulb air blower.
36. Replace the cryomicroscope stage lid to minimize dust ingress.
It is not necessary to tighten the lid completely.
37. If additional experiments are planned, reload the previously
saved temperature profile (Table 1) into the Linksys software
(see Note 97). When saving of the current video has completed
(see Note 95), repeat the above protocols starting with the
sample preparation procedure (Subheading 3.5).
3.8 Experiment
Shutdown
1. Set the stage temperature to a value in the range 30–40
C, by
modifying the active row of the temperature profile table in
Linksys (see Note 39); specify an indefinite hold (e.g., set
Time ¼ 99,999).
2. Maximize clearance between the BCS196 lid and the microscope objectives by using the coarse focus controls to lower the
stage (see Notes 49 and 50).
3. Detach the LNP purge tubing extension from the BCS196
front gas port by pushing the valve sleeve toward the stage
body until the valve clicks shut.
4. Disconnect the LNP liquid nitrogen withdrawal tubing from
the coolant exhaust pipe on the BCS196 stage (see Note 98).
5. Disconnect the dewar’s insulated capillary tubing from the
coolant inlet pipe on the BCS196 stage (see Notes 98 and 99).
6. Terminate control of the stage temperature, by clicking the
blue square button in the Temperature Control toolbar in the
Linksys interface to stop execution of the temperature profile.
7. Select “Disconnect” from the “File” menu; confirm that the
temperature display reads “No Comm.”
8. Exit the Linksys32 software, and then power down the liquid
nitrogen pump and the cryomicroscope system controller.
9. Unclasp the latches of the lid on the LNP dewar, and gently
remove the lid. If necessary, discard any leftover liquid nitrogen
using appropriate disposal methods.
10. Detach the LNP window tubing from the BCS196 stage lid,
and remove the metal clip from the end of the tubing.
11. Disconnect the LEMO connector from the socket on the left
side of the BCS196 stage body, by pulling it straight out.
12. When all image and videos from the experiments have been
saved, close the camera control software and shut down the
computer.
13. Power down the camera by disconnecting its power adapter.
14. Disconnect the data transfer cable and the trigger interface
cable from the camera.
High-Speed Video Cryomicroscopy
239