At the end of May, Bee and Chris traveled to Vancouver to attend the annual International Society for the Advancement of Cytometry (ISAC) Congress. Below are the highlights of the meeting which we would like to share with you.
Tuneable fluorophores: During the innovations session two companies showed off their concept of tuneable fluorophores. The idea being that you can alter the brightness of the fluorophore to match the expression level of the antigen being analysed. In theory, by doing this you can choose well separated fluorophores without worrying about brightness, allowing improved data resolution. Both use long chains of fluorescent molecules, where adjusting the length changes their brightness. The difference is in their construction; Phitonex uses DNA scaffolds to hold the molecules, whereas StabiLux use Boron Nitride nanotubes.
Activome: Katarzyna Groborz from Marcin Poręba’s group at the Wroclaw University of Technology discussed their concept of the activome (an –ome that sits on top of the genome, epigenome, transcriptome etc.) through the use of activity based probes directed at enzymes. These probes, when conjugated to metal isotopes for detection by mass cytometry (Helios and Hyperion systems), allows the current active metabolic processes to be probed in the cell.
Optogenetics: Is the process of using light to control biological processes in cells. There is a nice overview in Max-Planck Gesellshaft: https://www.mpg.de/18011/Optogenetics. During the innovation session Opto Biolabs showed us their flow cytometer add-on that allows real-time analysis if optogenetic processes on the flow cytometer. They are currently looking for trial partners (beta testers).
Microbubbles: Akadeum were showing off their novel cell isolation kits that use microbubbles to separate cells. The concept is the same as the Invitrogen, StemCell, and Miltenyi magnetic bead isolation kits, but using buoyant bubbles. They have developed a way to make these bubbles stable over long periods and to attach antibodies to them. This way you don’t require a magnet or column to separate your cells. There is a sample vial on Chris’ desk if you would like to see them.
opt-SNE: Anna Belkina talked about her preprint in bioRxiv detailing a method to optimise parameter choice when using tSNE dimensionality reduction, especially useful for larger datasets. The paper goes into some detail about the effect of various parameters and how to calculate when you have reached the “ideal” iteration. opt-SNE is available in the latest version of FlowJo and at this link: Omiq.
FAUST: There were two posters from the authors of the FAUST (full annotation using shape-constrained trees) algorithm for discovering and annotating populations. They take the direction of “standardizing and partitioning the space across samples and using the discovered phenotypes to match discovered cell populations across independent samples”. They subsequently used this to discover novel biomarkers in previously published studies. The preprint is here: https://www.biorxiv.org/content/10.1101/702118v1 and the code is here: https://github.com/RGLab/FAUST
metaCyto: Atul Butte from the University of California discussed his groups’ tool that allows the comparison and combination of mass and fluorescence cytometry data. metaCyto clusters both types of data together using silhouette scanning.
Berkley lights: The Beacon has been out for a while, but it has now be supplemented with a significantly cheaper lower throughput model called the Lightning, aimed at research labs. If you have not seen Berkley Light’s machines then watch this video now. They use light to move individual cells into pens, where they can grow, be stimulated, identified, and moved back out at the touch of a button.
REAlease antibodies: Miltenyi released these late last year. They are antibodies that can be fully removed from the cell after use. The idea being you can remove them after sorting so they don’t interfere with downstream processing and cell culture, or they can be easily removed for multiplexed sequential image analysis.
REAfinity antibodies: Miltenyi has also geared up in making high quality antibodies that offer superior lot-to-lot consistency and purity as compared to conventional mouse or rat monoclonal antibodies. They have been engineered to lack any background binding and also with the same human IgG1 isotype, eliminating the need to include multiple isotype controls during flow analysis.
Spectral analysers: Sony launched their new spectral analyser, the ID7000. This instrument uses 188 detectors to analyse the spectrum of a fluorophore to allow the separation of similar dyes that would not be possible on a traditional flow cytometer; such as GFP and YFP or APC and AF647. There are two spectral analysers currently on the market, the other being the CytTek Aurora.
Advancement in Imaging flow cytometry
High speed Imaging: Hideharu Mikami from Goda’s Lab has developed a novel way to increase the throughput of fluorescence imaging flow cytometry (FIFC) by virtual-freezing of flowing cells and spatiotemporal control of excitation beam illumination to allow image acquisition with a high throughput of ~10,000 cells/sec without compromising imaging sensitivity.
Image free ‘Imaging’ flow cytometry: Toru Imai from Thinkcyte Inc., has make a recent development and advancement for an ultrafast fluorescence ghost ‘imaging’ activated cell sorter (FiCS). This work is being carried out by integrating a microfluidic cell sorting platform with high throughput cell classification based on supervised machine learning algorithm.
High speed Imaging Image free ‘Imaging’ flow cytometry