Flow Cytometry Data is Beautiful

So, why not make your latest flow analysis into a work of art? Read on for info regarding the "Science in Art" exhibit, or visit uchisciart.org for more details.

Science in Art 2008 Issues Call for Art
Science in Art, a juried art exhibit that features art from scientist-artists from The University of Chicago, Argonne and Fermi National Laboratories and Chicago artists whose subject is science, is accepting art submissions for the Science in Art exhibit 2008 and will accept submissions through Friday, August 22, 2008. The exhibit was developed in response to the need for educating the public about the process, challenges and benefits of science and technology.
Science in Art uses art as a vehicle for creating connections between scientists and non-scientists. As science and technology advance the need for public awareness and understanding increases. The success of science depends largely on public support, therefore communication between scientists and the public is critical. This exhibit will help to translate the language of science for the general public and to push forward the boundaries of human enlightenment. Moreover, it will highlight cross-disciplinary connections in the development, expression and exploration of novel ideas.
Art media accepted for submission include photography, drawing, painting, sculpture, mixed-media, videography and music. Science in Art organizers and a distinguished panel of jurors will select art for the exhibition based on three criteria: (i) how the artwork relates (abstractly or literally) to science (ii) quality of execution (iii) How the artwork goes beyond the mere depiction of the scientific phenomenon.
The Science in Art exhibit will take place at The University of Chicago Gordon Center for Integrative Science (GCIS) Oct. 10-Dec. 13 with an accompanying opening reception on Oct. 10.
Complete entry information, including an application for submitting art, can be found at: uchisciart.org, or by contacting Rebecca Ayers (rayers@uchicago.edu; (608)345-1321.

'Not all DI water are alike' or 'Why the Aria Broke.'

Symptoms: Anything we put through the FACSAria, came out dead. Not like exploded, obliterated dead, just permeable to trypan blue, not-able-to grow, dead. Secondly, tandems looked weird. For example, PE-Cy7 would have a PE-Cy7 positive population, but then it would also have a PE positive population, as if the Cy7 portion of the tandem was getting quenched. However, the PECy7 antibody was fine since it looked normal on the MoFlo.

Troubleshooting: Obviously, the 1st thing to check is the buffer. So, I collected some PBS exiting the Aria's flow cell, mixed that buffer with some cells, and ran them on the instrument and checked them under the microscope. They were fine, suggesting the buffer itself was not killing the cells. I then turned to the (HPLC) valve that controls sample uptake. I disconnected the sample line, after the valve, but before the flow cell, collected some cells, and then reran them. They were fine again, suggesting that it's not the HPLC valve that is killing the cells. So, it seems like the cells have to pass through the flow cell in order to die. On the flow cell, there are wires attached to provide the charging of the stream (required for sorting). So, I disconnected all the charging wires and tried again...still dead. Maybe the lasers are frying the cells. Turned the lasers off...still dead. What to do next?

Placing a volt meter on the metal portions of the flow cell, we discovered a 0.5V charge present. So, perhaps if we grounded the flow cell with an external copper wire attached to a grounding source (in this case, an electrical box behind the Aria) we could keep the cells alive...no luck!

Then, it hit me like a ton of bricks. For about a week now, our Millipore Biocel Water Deionizer was down, so I was getting deionized water from the Monoclonal Facility. Now, the monoclonal facility uses this water for all their tissue culture work, so it is definitely a good source of H2O. The output of water was 'suppose' to be comparable. But then I thought, if the problem really is an electrostatic charge in the flow cell, and, if the new water source is not deionized to the same degree as our original water source, the electrostatic charge could be passing through the buffer, directly to the cells and basically shocking them. Pretty much like an aggressive electroporation.

All the pieces were coming together. It makes sense. This could also explain the PECy7 issue. The extra electrostatic charge was messing with the fluorescence of the fluorochromes as well. So, the obvious next step was to use water from a Biocel Millipore unit. As soon as I did that, all was fine.

So, in summary, buffer made with water from this specific Millipore system, passing through the flow cell on the Aria, with cells present shocked the cells and killed them. Buffer made with water from the Millipore Biocel system..fine. I never found out from BD if that electrostatic charge is suppose to be there or not, but it seems like properly deionized water is required to protect the cells from this charge.

All is well now. Happy sorting!

The new flow cytometry Zune?

So, not really flow cytometry news, but a mockup of the soon-to-be-released special edition, "Joy Division" zune struck me as oddly familiar. The etching on the back. Anyone see a bunch of overlaid flow histograms ala Flowjo? Flow cytometry is now influencing pop culture, or at least pop culture from the late 1970s!

University of Chicago Cancer Research Center achieves Comprehensive Status

For those of you who are not aware of the University of Chicago Cancer Research Center (UCCRC) on campus, I'll share a bit of good news. First of all, the Cancer Research Center Support Grant has been renewed for 5 years, and secondly, the UCCRC has achieved Comprehensive status. This is sort of a big deal, considering in the entire country, there are only 41 Comprehensive Cancer Research Centers, and in Illinois, there are only 2 (the other one being the purple people downtown). Being a Comprehensive Cancer Research Center basically means we've got really great researchers and support staff working on attacking malignant diseases, or as the NCI likes to say, "Comprehensive Cancer Research Centers are characterized by scientific excellence and the capability to integrate a diversity of research approaches to focus on the problem of cancer."

So, what does all this have to do with flow cytometry? Well, a big part of the UCCRC grant was the availability and access UCCRC members have in terms cutting-edge technology and technical expertise. A big chunk of the funding goes towards operational costs of core facilities and subsidized usage fees for member investigators. The flow cytometry facility, having a large number of cancer research investigators has been the recipient of such funding for over a decade. Over 90% of the usage of the facility is performed by members of the UCCRC, so we are very much an integral part of the UCCRC and its newly achieved Comprehensive status. More info on the UCCRC can be found online at http://uccrc.uchicago.edu or if you'd like more info on the NCI Cancer Research Center Program, you can visit http://cancercenters.cancer.gov. Congratulations to all involved!

ISAC Wrap-up Part 2

Along the same vein as the previous post, the Education committee presented an outline of their plans to bring flow cytometry education to the general scientific public. In an attempt to standardize the information being presented to users of flow cytometry, the Education committee has decided to generate a basic flow cytometry course aimed at flow novices. The course would initially be offered as a tutorial tacked on to the front end of other disciplines' meetings, who have used flow cytometry in the past. From there, plans to make it available as an online course administered through ISAC were discussed. Lastly, it was proposed that this course become the template for training used by core facility directors in training their user base. The Education committee still seems to be developing these ideas and reworking their strategies, so it may be awhile yet until we see anything concrete. Don't know yet if there will be an official "accreditation" process for this type of course, but if and when it's possible, we'll be sure to take a good look at the material to see how we can integrate it into our educational process.

ISAC Wrap-up Part 1

Planning to submit a paper to J. Exp. Med. anytime soon? Well then you'll want to pay attention to the new guidelines generated by the Data Presentation Standards Committee and adopted by J. Exp. Med. What the cytometry field has been noticing for quite a while is the amount of publications in high-end journals with really poor flow cytometry figures. It's not that the data is bad, or doesn't support the authors conclusions, it's just that the figure is annotated so poorly, and the methods written so vaguely that it's nearly impossible for anyone to try and repeat complicated analyses. Being the premier flow cytometry authority, ISAC has taken it upon itself to lay down some guidelines to assist authors in properly presenting the necessary information to describe their flow cytometric data. A member of the Data Standards Committee, Dr. Mario Roederer, shared the conclusions of the committee this week at the ISAC Congress. Here are a few examples from the guidelines. Once a comprehensive list is created, I'll post them for the group to see.

Author must list the instrument(s) used, the laser(s) used for excitation, and the filter(s) used for emission. This will be done in an excel template that will be available from ISAC or J. Exp. Med.

Axis on plots must have the reagent and fluorescence labelled (ex. CD3-FITC).

The number of events in a plot must be displayed in the plot or figure legend.

The frequency of populations must be displayed on the plot or in a table

Graph-types should be used consistently throughout.

You MUST show the entire gating tree for your figure as an example. Back-gating analysis is the best way to show this. This can (and probably should) be in the supplemental data portion of the manuscript.

You must also state how you drew your gates. For example, did you draw them based on unstained cells, or an isotype control, or and FMO control, or did you just subjectively draw it around a population.

These are a few of the guidelines that will be implemented soon. J. Exp. Med. has decided to adopt them sooner rather than later, but we anticipate more and more journals will begin to require this information. If you have any questions, or need help filling in the appropriate information, the flow can certainly help. We will try to provide filled out examples for each of the instruments so that you can simply modify them for your use. We'll provide more info as it becomes available.

Greetings from Budapest

This year's ISAC meeting, held in Budapest, Hungary is well under way. I'll post some interesting tidbits of info as the meeting progresses. One interesting tidbit is that ISAC has changed it's name effective immediately. It will still retain the acronym ISAC, but the last two letters have been slightly modified. Previously, we were known as the International Society for Analytical Cytology. That name, analytical cytology, doesn't really fit with what the society does, so it was voted on and it was decided the new name will be "International Society for Advancement of Cytometry" Cytometry is what we do, that is, measure cells. Also, our goal is to advance the field of cytometry, so the name makes much more sense. More later.