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Principles & Protocols

Protocols - Using the Bal-Tec High Pressure Freezer

Jump to Principles

• If you are having problems turn the HPF off and restart
• Download MSWord version of these procedures: HPF-procedures.doc

START-UP

• Fill water recirculator
• Turn on 2 power switches 1hr before use (objecthead temperature must read 40 b4 use)
• Fill with 2-propanol
• Turn on N2 tank (meter should read between 4-5 bar)
• Purge system with N2 for 30-40 min (air heater button)
• Turn on LN2 tank
• Push START
• Push NITROGEN
  • If no noise from LN2, tap gently on solenoid valve inside left side
• Wait until red NITROGEN light goes out
• Push DRIVE IN
• Wait until drive in light goes out (pressure should increase to ~200 bar)
• Push AUTO (pressure should increase to 310 bar and subsequently cycle 295-310)
• Record freezing parameters with measuring holder before commencing
  • Make sure planchette is loaded in measuring holder
  • Pressure Maintenance should remain >0.5 sec, or grease/change o-ring
  • Use laptop and Matlab to record analogue trace of pressue and temperature
  • Save pdf of trace on laptop

CHANGE LN2 TANK DURING RUN

• Push STOP
• Warm up LN2 connection with heat gun
• Change tank quickly, especially when humid
• Push START
• Push NITROGEN
• Wait until red NITROGEN light goes out
• Push AUTO (drive in not necessary)

SHUTDOWN

• Push STOP
• Turn off LN2 tank
• Turn off N2 tank
• Release pressure at 2-propanol inlet
• Turn off key switch
• Leave on Mains switch this will continue recirculating the water for 10 hours before shutdown
  • make sure water bath is full of water
• Remove holder from Objective head and put on black plastic cap

Visit Cryo EM website at http://www.nysbc.org/facilities/CEM

-- DavidStokes - 18 Mar 2005