Start from a clear question
First things first
- What is the best strategy to quench metabolism?
- What is the best way to extract my samples?
- What is the best way to store my samples?
- MS performance degrade with time: how long can I run my sample list before cleaning
Controlling analytical variability
- Block your experimental design on the analytical batch
- Reduce as much as possible the length of the chromatographic run
- Measure the analytical variability
Your silver bullet: Quality Contro Samples (QC)
Reference samples which are regularly injected along the sequence.
They measure analytical stability because the sample itself is the same.
QC samples should be representative of the chamical complexity of the samples
Pooled samples are often the preferred choice
A fairly good sample list
- The usefulness of blanks is disputed
- The first repeated QCs are use to allow the column to equilibrate
- One could add QC dilutions
- Also the design of the sequence can be validated
Other relevant points of discussion
- Standards and blanks
- Analytical batches and batch effect
- Randomization, analytical batches and large studies in clinical settings
DDA and Fragmentation
Experiment combining full scan and fragmentation experiment are quite popular, because fragmentation patterns are needed for metabolite annotation and for setting-up quantitative methods
Use wisely your time
The more experiments I do, the worst is my estimate of the peak area …
- plan different runs for FS and DDA
- fragment only ion of interest
- strike a balance between chromatography, instrument speed and number of experiments