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Technology

Just the FACS, man.

Many scientific institutes have a need for core facilities to process samples in a ‘cheap’ and efficient way. These centralised units have a big advantage over separate groups purchasing expensive pieces of equipment: they can pool financial resources and employ managers and operators with a high level of technical expertise to get the best possible results. Flow cytometry or FACS (fluorescence-activated cell sorting) is an important scientific resource and Kevin Clark, a senior sort operator in the WIMM, explains the critical role that it plays in current research.

Flow cytometry is a technique that allows us to identify different types of cells based on their physical and chemical make up. The WIMM has twelve highly specialised pieces of FACS equipment (cytometers) that serve the needs of some 300 scientists in one core facility.

Image credit: Kevin Clark (2014)

Image credit: Kevin Clark (2014)

Imagine you’re watching thousands of cars passing by on a road and you’re noting down the colour of the car, as well as the colour of the driver’s shirt, hair and eyes and you were only interested in green cars where the driver had a red shirt, dark hair and blue eyes. Flow cytometry allows us to make that kind of selection with cells at incredibly high speed.

Cells at different stages of development have different structures on their surface. These structures can be labeled with different coloured dyes (fluorochromes) that can be turned on when passing through a laser beam. FACS allows us to identify and collect cells for further analysis based on the presence or absence of different combinations of these colours.

The cells are suspended in a stream of buffer solution and pass one at a time through different laser beams to make them fluoresce or glow. The cytometer can identify the different colours and allow us to separate out the cells that we’re most interested in, based on that colour profile.

The cells are extracted from the stream using high voltage (it doesn’t kill them!) and they drop in to a collection tube. We can interrogate around 25,000 cells per second whilst checking up to 15 different colours and sorting up to four separate populations at once.

My role is primarily as a sort operator to perform cell sorts for scientists in the Institute, but also to advise on data and experimental setup, give technical assistance, teach people how to use the analysers (which allow them to run samples themselves) and do simple general maintenance on the multiple sorters and analysers we have.

Simple eh?! Well, no.

Image credit: Kevin Clark (2014)

Image credit: Kevin Clark (2014)

Sorting itself is challenging. You need to know the full ins and outs of how the cytometer works in order to get the best out of it for the user. Control samples are run first to assess the baseline of the fluorescence in all of the colours you’re using and to check that the staining of the fluorescent proteins used has worked correctly.

We also need to take into account how similar the different colours used are: if you try to use colours that are too similar, this can cause serious problems for the cytometer. For example a light green and a dark green car are different even though both are green!

We can then determine the percentages of the cell populations in the sample, the existence or absence of a population of interest and decide the cells we want to collect. Sorts can take anything from 5 minutes to 5 hours depending on the number of cells being processed and the complexity of the sort. An iPod full of quality music is essential!

So that is the life of a flow cytometrist. I was once asked, how could you have a machine with lasers and NOT try setting fire to things?*

Hey, just the FACS, man. Just the FACS.

*Besides which it breaks numerous health and safety rules…..unfortunately!

Post edited by Bryony Graham, Gemma Swiers and Paul Sopp.

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  1. Pingback: How students see scientists: Part IV | The WIMM blog - March 10, 2015

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