The Art and Science of Flow Cytometry

Life in a Parallel Universe

John DaleyJohn DaleyDirector Director

Flow CytometryFlow CytometryHematologic NeoPlasiaHematologic NeoPlasia

Dana-Farber Cancer InstituteDana-Farber Cancer Institute

Definitions

ScienceThe classification of knowledge gained by observation

and experimentation

TRUTH

TR

TH

ARTThe seeking for an

Ideal truth

What makes a Good Sort

• Purity• Recovery• Integrity

What makes a Good Sorter• Reliability• Reproducibility• Responsiveness• Stability

The Other Part of the Picture• People

Operator:Investigator:

Supporters:Places: Environment makes it work

Critical Mass• Multi application exposure • Large user base generates support and application

development• Equilibrium dynamics coexist between instrument,

operators, support, and users• Interaction dynamics between Experiment, cells ,

instrument, researchers, and operators• Active Focus on priority at hand: Sort in Progress• Not too Big not too small

When things go Right• Keep going, don’t look back• If it works: Duplicate, duplicate, duplicate• Be on the lookout for hidden hisses and the

sounds of sparks• Stay calm• Keep a mindful eye on the relevant features• Trust your senses! • Be humble

When things go not so Right• Stay calm• Think what are the components of the issue• Try the easiest thing first• Remember past experiences • Employ back up system if possible• Start from beginning• Replace the o ring• Reboot the computer

The Rituals of Flow• Startup• Shutdown • Keeping control• Sort Set up• Active Sort• Post Sort• Sterilization

When the Sort works well• Everybody is happy• Still need to follow cells down the data path• The Proof is in the Pudding• Faith in the System, Operator and Protocol• Creates confidence for the next experiment• Proves Preparation was worth it

How we did it :1996-2007• Open door policy as much as possible• Educate ourselves and users about the science and

technology• Segregate analysis and sorting• Maintain as sterile as an environment as possible• Tried to keep equipment as up to date as possible• Constant form is maintained by constant growth• Created a long range Plan• Created a concept of supported self service system• Always had a back up instrument system in place• Always placed experiments on appropriate instruments • Learned from our mistakes in trusting others

Time for Expansion• 1999: need for high speed and space

reorganization• Room consolidation : infrastructure• Due diligence for instrument best suited for

our needs: • Critical mass established: need created a

confidence that more powerful equipment would be utilized and self supported

Facility Structure• Organize User priority• Create a common information system: web

page/ scheduler• Expand tentacles of facility via extended

analysis• Creating an internal accounting system that

could serve as a check in rectifying discrepancies between vendors and administration

Don’t be an Operator Be a Scientist

• Educate• Information resource• Never say No (most of the time)• Try not to get taken advantage of• Mutual Respect is key• Above all Maintain a sense of Humor

The Good the Bad and the Ugly• When private flow went core• When operators were considered paying

employees• When the bottom line of cost recovery

creates an atmosphere of greed• Is flow a business or a science?

How to make a sort work• It’s in the set up• What's needed: operator decides, let user be

part of the design , operator maintain transparency

• Always do a final pre check right before liftoff• Transfer a sense of calm to the responsible

parties• Be part of the sort