Automatic Deposition of CHCA Matrix for MALDI Analysis of Lipids
Elizabeth Neumann, Carrie Romer, Jamie Allen, Jeff Spraggins
Abstract
Scope:
To describe the procedure for spraying tissue sections with CHCA for lipids. CHCA is used as a safer matrix to ship coated slides to collaborators.
Expected Outcome:
Slides should be coated with CHCA. Tissue sections should be imaged within 24 hours of matrix application.
Steps
Autofluorescence Scan
Remove slides from freezer and place in desiccator for 0h 30m 0s .
Scan for autofluorescence on Zeiss Axio Scanner.
TM Sprayer Setup
Change nitrogen to 6 psi and turn on sprayer.
Open software and set nozzle temperature to 85°C
Change LC solvent to 70% methanol at 0.1 mL/min.
“Load” sample loop with 5mL methanol.
Switch sprayer to “Inject” and spray for 2 minutes.
“Load” 6mL of matrix solution into sprayer loop.
Switch sprayer to “Inject” and spray for ~1 minute. Use slide to check that matrix is flowing.
Tape slide(s) to the top of the stage and adjust software scanning area.
Set method in software:
700 mm/min nozzle velocity
2 mm track spacing
0.1 mL/min flow rate
CC Pattern
2 L/min flow rate
8 passes
40mm nozzle height
60 second drying time
Save method and make sure it is highlighted.
Under Cycle, click “Start,” then click “Continue.”
When finished, remove slide and place in slide box.
Cleanup
Set TM sprayer temperature to 30°C .
Switch A1 solvent line to 100% ACN and set flow rate to 0.5 mL/min.
“Load” loop with 20mL acetonitrile and “Inject” loop for 0h 10m 0s .
Spray tip of nozzle with MEOH.
Log the pressure at your flow rate post cleaning. It must be within 3-4 psi of the starting pressure; if it is not, the cleaning procedure must be repeated.
Change the HLPC flow rate to 0.05 mL/min.
Spray nozzle with methanol.
Remove the foil and wipe stage down with methanol.
Replace wypall and bench diapers.
Place all syringes in the biohazard container and empty the solvent waste.
Shut down the software and turn off the TM Sprayer.
Shut the nitrogen off.
