Controlling or at least
Variability In a core facility environment
• Increased variability = decreased power
• Power = probability of find an effect that is there
• You can fight this by increasing the sample size but
often it is much cheaper to decrease variability
Common sources of
• Sample preparation
• Data Analysis
What is possible in a core
• Almost no one who sends me samples has enough
money to measure variability or wants to pay for it
• What are the best ways to communicate these
issues to customers?
• How do you know what your variability is if there are
no resources to measure it?
• How do you measure variability when you have a
large number of different types experiments?
• How much QC do you bundle into your costs if you
have to charge people
Some issues I routinely
• Analyzing samples over months at a time….
• Sample preparation of Plant tissue may be
completely different than human cells or Plasma, or
Milk In terms of how consistently you can prepare it
• How do I know how consistently I can prepare a
• Often I have no control over how the sample is
prepared. How do I deal with that?
Common ways to decrease variability
during sample prep
• Process all samples on the same day by the same
o Person can still get tired or make mistakes…Variability may not be
consistent beginning to end
o May not be possible
• Use Robotics for part of the sample prep
o Many things still cannot be done well by robots
• In gel digestion of proteins is not optimal
• Decrease the things you do to a sample
o Fractionation, precipitation, SPE
• Label proteins or peptides upstream and multiplex
Common sources of variability you may not be
• Pipetting errors
o Can be vary large for small volumes
• Eppendorf 2 ul = 12% Systemic 6% random error
o Hard to get tight cv’s on your spiked peptides
• Variability due to SPE material lots
o The SPE material you use today may not be the same the next time you
• Variability due to software
o manual integration
• Are empirical Null’s a good way to measure
Is peptide or protein
fractionation worth it?
• Does the fractionation kill your power?
• Is it better not to fractionate ?
• What is the least variable fractionation method for
• How do you measure the variability your
Example method to
• From Chris Becker (Proteometrics)
are pooled before
replicates are run
are run individually