Quanta 450 ESEM – Operating Manual (Revised with Safety Warnings & Tips)
Important: You must arrange with FEMR staff before operating the microscope in ESEM mode with the cooling (Peltier) stage.
Safety Warnings for the Quanta 450 ESEM
⚠️ High‑Voltage and Electrical Safety
- The microscope operates at high voltage—never touch internal electrical components, detector contacts, or cables unless instructed by FEMR staff.
- Do NOT plug or unplug detectors with the beam on or while the system is actively pumping.
- Ensure all cables are fully seated; loose connections may cause arcing or detector damage.
⚠️ Vacuum and Pressure Safety
- Never open the chamber door until the system reads VENTED; pressurized gas release can cause injury or damage.
- Do not obstruct or cover vacuum ports, vent lines, or pumping pathways.
- Sudden pressure changes can cause sample material to eject—secure samples properly.
⚠️ Moisture and Condensation Hazards
- Moisture inside the chamber is very dangerous for detectors and the pole piece.
- Never exceed 80% humidity, and never bring the working distance below 8 mm.
- Be cautious about condensation and water leaks, as they can seriously damage the instrument.
- If you see condensation forming on the stage, detectors, or chamber walls, stop immediately and call FEMR staff.
- Do not insert visibly wet or dripping samples.
⚠️ Cooling and Water System Safety
- Never exceed the recommended minimum working distance (8 mm)—moisture can enter the pole piece, causing permanent damage.
- Use only distilled water in the chiller system—other water types can corrode components.
- Always check for water leaks after connecting the water lines. Stop immediately if leaks are detected.
- Never run the Peltier stage without the water chiller switched on.
- The cooling lines and stage may become very cold—avoid direct skin contact.
⚠️ Sample Safety
- Biological samples must be handled using PPE (gloves, lab coat, eye protection).
- Unfixed biological material can contaminate the chamber—clean thoroughly with 70% ethanol afterward.
- Do not load samples that exceed the height of the sample holder; they may contact the pole piece during pumping.
⚠️ Detector Handling
- Do not touch internal components (detectors, pole piece, stage hardware) unless explicitly instructed.
- Handle detectors gently — Detectors (SED, GSED, GBSD) are fragile (and expensive)—never pull on cables, connectors, or sensor heads.
- Place removed detectors only on padded, clean surfaces.
⚠️ Stage Movement Hazards
- Keep hands clear of the stage and column when moving the stage.
- Never raise the stage while looking away from the chamber image—crashes can permanently misalign the column.
- When switching from the Peltier stage back to a standard holder, ensure the stage is fully lowered before inserting any hardware.
⚠️ Chemical and Cleaning Safety
- Only use approved cleaning agents (70% ethanol, lens tissue where appropriate).
- Do not use acetone or strong solvents inside the chamber—they can damage seals and coatings.
⚠️ Thermal Safety
- Cooled components can cause frostbite‑like burns—avoid direct contact.
- Heated samples (if used) can cause burns—use tools, not fingers.
⚠️ Emergency Shutdown Conditions
Immediately stop imaging and call FEMR staff if you observe:
- Unusual noises from pumps or chiller
- Water or condensation near the pole piece
- Sudden pressure spikes or failure to reach vacuum
- Smell of burning, ozone, or overheated components
- Detector error warnings during ESEM operation
1. Prepare the Caron Water Chiller
- Switch on the Caron water chiller behind the Quanta 450.
- Confirm the temperature setpoint is 10 °C.
- Fill reservoir with distilled water only (⚠️ never deionized).
Safety Tip:
👉 If the chiller tank runs dry, the Peltier stage can overheat—always check water level before activation.
2. Configure Initial Microscope Settings
- Set Accelerating Voltage to 15 kV and Spot Size to 3.5.
- Vent the chamber and open the door.
Safety Warning:
⚠️ Wait for complete venting before opening the chamber door to avoid pressure‑related hazards.
3. Install the Peltier Cooling Stage
- Remove the decelerating stage if installed.
- Position the Peltier stage with water lines and ribbon cable facing backward.
- Lightly tighten the helicoil center set screw.
Safety Tip:
👉 Do not overtighten the helicoil screw—it can strip easily.
4. Connect Cooling and Control Lines (Staff only)
- Remove the water plugs inside the chamber.
- Insert the water lines and connect the vertical 9‑pin ribbon cable.
- Verify that the COOLING button activates.
Safety Warning:
⚠️ Ensure the sample fits entirely within the holder—protruding samples may contact the pole piece during pumping.
5. Prime the Water Valve System
- Power on the Water Valve Box (left of column).
- An alarm will sound—press & hold the GREEN button until water flows smoothly with no air bubbles.
- Check for any leaks.
Critical Warning:
⚠️ Immediately stop if water leaks inside the chamber—alert staff before continuing.
6. Install the Appropriate Detector (staff only)
- Carefully remove the Wide‑Angle SED.
- Install the GSED or GBSD and clip the axial endpiece to the pole piece.
- Close the chamber and press VACUUM.
Safety Tip:
👉 Hold detectors by their housings—not their cables or sensor heads.
7. Cooling & Humidity Control
- Activate cooling under the Temperature Control tab.
- Open the Humidity tab to view the Triple‑Point Graph.
- Recommended starting point:
- Temp: 5 °C
- Pressure: 5.5 Torr
→ Produces ~75–78% humidity (stable, moderate moisture)
Safety Warning:
⚠️ High humidity (>80%) can cause water droplets to form on internal components—risking electrical and vacuum damage.
8. Adjust Temperature & Pressure
- Input desired Pressure in the main menu.
- Adjust Temperature → press GO TO.
- Monitor stabilization under the Humidity tab.
Tip:
👉 Make small adjustments—ESEM humidity is highly sensitive and changes rapidly.
9. Pre‑Imaging Setup
- Press BEAM ON.
- Center the stage.
- Raise stage until Peltier stage edge is just above the yellow 10 mm line.
Critical Warning:
⚠️ Never approach closer than 8 mm to the pole piece—moisture can be pulled into the column.
- At 100×, increase to 400× and use Auto Brightness/Contrast.
- Focus on the bottom of the sample holder.
Safety Tip:
👉 Lens alignment should be done now—it is difficult in high‑humidity ESEM mode.
10. Load the Sample
- Turn BEAM OFF.
- Vent the chamber fully.
- Pre‑cool the sample when possible.
- Load onto stage; optionally add water to the four dimples.
- Close chamber and press PUMP.
Biological Safety Warning:
⚠️ Wear gloves and avoid introducing contamination inside the chamber.
11. Begin Imaging
- Press BEAM ON and start at 400×.
- Find a feature to focus on.
- Link stage height.
- Set WD ≥ 8.00 mm.
Safety Tip:
👉 Do not adjust WD below 8.00 mm even if the sample appears flat—risk of condensation on the pole piece increases at close distances.
12. End-of-Session Procedures
- Restore all settings to defaults:
- Cooling Off
- Pref:Scan:ESEM default
- Stage lowered fully
- WD fully down
- Drain water lines; disconnect water and control cables.
- Remove the Peltier stage, reinstall the single-stub holder, and reinstall the SED (Staff only).
Safety Warning:
⚠️ Water remaining in lines can backflow into the microscope—ensure they are fully drained before disconnecting.
13. Cleaning & Decontamination
- Clean all holders, counters, and nearby surfaces.
- Remove all particles, tubes, or biological debris.
- After unfixed biological samples, wipe with 70% ethanol.
Tip:
👉 Allow surfaces to air-dry fully before closing the chamber to avoid moisture spikes during the next session.
14. Final Safety Note
Before inserting the standard specimen holder, always fully lower the stage after removing the Peltier stage to prevent collision with the pole piece.