Monday, November 21, 2011

Options for Flow Cytometry Training - FloCyte Review

Flow Cytometry (FCM) isn't the easiest technique to learn.  It actually takes quite a while to master both the hardware and software components to sample acquisition and data analysis - let alone the applications utilizing the aforementioned instrumentation.  For many users of flow (in an academic setting) their first encounter with FCM is likely through a core facility, whereby they'll receive some instruction on how to operate an instrument and then how to analyze the data they collected.  The type and quality of this training varies greatly.  Some institutions I'm familiar with have multi-day courses with wet lab sessions and hands-on instrument time, while others attempt to provide a theoretical base and then do a bit of hand-holding for a few sessions.  The success a user may achieve greatly depends on his or her resourcefulness and overall aptitude for technology.  Some people pick it up quickly; others struggle for years.  I will say that training users in a busy core facility is a huge drain of time and resources.  In our core, for example we basically have an entire F.T.E. just providing training and consultation, so I'm sure that in smaller cores, where it's just one or two people, training has to be an even greater burden.  The question then becomes, how are we to provide the necessary training and attention our users require with the limited time and personnel resources characteristic of a core facility?

There aren't too many options.  Before I jump into an assessment of the FloCyte courses (which is the whole point of this post) let me briefly highlight other possibilities.  FYI, I've personally attended all 3 types of training sessions and have viewed all the resources in #4.  

1.  The Annual Course in Flow Cytometry - This weeklong course alternates between Los Alamos National Labs (or the University of New Mexico) and Bowdoin College in Brunswick, ME.  It is really geared towards users of the technology who already have a basic understanding of the technology.  Also, it focuses on the applications of flow cytometry rather than operation of a flow cytometer, however numerous sections also delve into the hardware components.  There's a pretty cool lab where you can assemble your very own (fairly crude) cytometer.  The cost of the course is about $1800, which includes dorm-style accommodations and meals (transportation is not included).   

2.  Vendor-specific instrument/software training - Most vendors will provide training for their hardware and associated software.  When you purchase an instrument, you might get some free training included with the purchase, but additional training is going to cost you.  As you'd expect, the training is geared towards the operation of that vendor's hardware.  If you were using multiple cytometers from different vendors, this obviously wouldn't be ideal, but if you were using a single platform it might be a good option.  The vendor training will also include some of the basics of cytometry, but again, it will be skewed towards their instruments, their reagents, and their idea of the technology.  It's also pretty expensive, sometimes as much as $2500 per person.

3. Training courses at meeting - Typically when you go to some of the bigger conferences they'll have some workshops on FCM.  Certainly at the CYTO meetings you'll have the opportunity to attend training sessions on various topics.  Also, some of the immunology focused scientific meetings will have some FCM training associated with them (for example, the AIC meeting in Chicago).  Cost for this training is variable, however it's usually limited to conference attendees, so unless you were already planning to attend the conference, it might be really expensive. 

4.  Online utilities - There is quite a lot of information freely available on the web.  You can certainly start at the Purdue University Cytometry Laboratory web site, where there are a bunch of powerpoint slides, movies, and resources freely available.  In addition, companies such as Becton Dickinson, Life Technologies, and Beckman Coulter offer overviews of flow cytometry and flow cytometer technology.  Note that the above links are linked directly to the company's training/support page with the intended materials.  Although these online utilities are readily available and free, you lose the benefit of asking questions and interacting with people who can tailor the training to your specific needs.  

So now, I'll walk you through my experience with the FloCyte Training course offered by FloCyte Services.  I attended the Comprehensive Training Course from 11/15/11 - 11/17/11 held at Spherotech, Inc.  I won't bother taking up space here to give you the rundown of the company and the mission of the training courses.  You can read all about it here.  However, I will note that I attended the Comprehensive training course, which is designed for novice users of flow.  You can see the course curriculum here.

Day 1, as you'd expect, goes over the basic components of flow cytometry.  This is done is a pretty common fashion, and anyone who's gone through the powerpoint slides on the Purdue University Cytometry Laboratory web site will recognize the format.  4-components, Fluidics, Optics, Electronics, and Data Analysis.  All the standard material you'd expect to be here is here.  There was however at least one pretty critical omission - multi-laser systems, laser delays, and how fluorescence emission is spatially separated.  I know this was briefly mentioned during one of the sections, but there was no figure, no reiteration of how it's possible to look at two colors with the exact same emission simultaneously because they're excited by spatially separated laser beams (e.g. PECy7 and APCCy7).  When we broke into small groups to take a look at some of the hardware, I spent most of the time explaining to my other group members how this works.  They were very confused.  The graphics used to talk about emission filtering where all systems like a FACScan or FACSCalibur, which don't have spatially separated beams, and all the light goes through the same "pinhole".  Also on day 1, we finished up with a mathematical explanation of compensation, which went horribly wrong.  The math is complicated and it's probably not something basic users need to understand in order to compensate their data correctly (or, should I say, let FlowJo compensate their data correctly).  Lastly, there was no mention of 1 very critical component to flow cytometry, Quality Assurance and Quality Control.  In all, the basics were handled just fine.  I will say, though, that it seemed to move pretty slow.  I think for the amount of information covered in that first day, it could've have been condensed into a half day.  For example, I feel like the flow basics class given at UCFlow is comparable in it's scope but is completed in about 1.5 -2 hours.  

Day 2 brought in a plethora of applications and tried to reinforce some of the concepts from day 1 while explaining how those concepts effect how you think about the applications.  I think this way of presenting the information is really good.  When we're talking about immunophenotyping, we're also talking about compensation, background due to fluorescence overlap, non-specific binding, etc...   When we're talking about cell cycle, we're also looking at doublet discrimination, coincidence, sample core size, etc...  Here we also start tackling the necessity of controls, including comp controls and the always popular FMO controls.  My big issues with this section solely revolved around the figures.  Many of the figures were at best poor representations of the idea being put forth and at worst blatantly misleading.  This was especially noteworthy in regards to an explanation of biexponential display transformation.  In another instance, the instructors were driving home the idea of how we are to never use quadrants to perform gating on our plots and the very next slide describing FMO controls was filled with quadrants used as gating.  A bit contradictory.

Day 3 was all about stats and panel design.  The stats part was very straight-forward and pretty easy to follow.  The panel design section was good, and covered many of the issues that arise when trying to put together a multicolor panel.  There was an introduction to a utility from Treestar called Fluorish (which I'm not going to complain about because I like it)  however there wasn't any real mention or demonstration of other available utilities like Chromocyte and CytoGenie.  Also, we spent some time going through some data analysis strategies using FlowJo.

The cost for the 3-day Comprehensive course is $700.  The beauty of the course is that it's brought to you (either your institution can host it, or it is hosted nearby) so you don't have to factor in airfare or hotel costs.  But, you'll have to remember that you're getting a comprehensive theoretical overview of flow cytometry, you are not learning how to operate your specific cytometer.  So, if you didn't have a core facility around to show you how to open up FACSDiVa and adjust voltages on your LSRII, you'd still be pretty clueless on how to run your first FCM experiment.  Another positive about the training is that it is modular such that you can attend just days 1 and 2, or just 2 and 3, or even just day 3.  That way if you have some basic knowledge already, you can skip day 1 and just attend days 2 and 3. Lastly, I'll mention that there are a bunch of other, more advanced courses available outside the comprehensive course, including a multicolor compensation course, a course on "phosflow" assays, and even clinical flow cytometry.  

The instructors are well-respected flow cytometry professionals with years of experience under their belts.  They presented most of the material in a clear and concise way.  There was, at times, some confusion regarding what a figure was trying to describe, but this was due to the fact that the slides were recently re-done and the instructors were not 100% comfortable with them.  I feel like I want to give them a pass on that, but then again, I did pay $700 on this course and expected a very polished delivery.  All things considered, they did an excellent job.

I could see this working in a couple of ways.  1.  You get some initial training on how to operate your cytometer from your core facility and then attend days 2 and 3 of the comprehensive course.  2.  You could attend the entire comprehensive course and then go through the specific instrument training given by your core facility.  3.  Get trained by your core, start running experiments, and then jump in on one of the advanced courses offered by FloCyte.  If you're not fortunate enough to have the support of a core facility, then this makes the FloCyte courses even more attractive.  Relying on them for the basic theoretical training, and then the instrument vendor for training on the actual equipment is probably your best bet.