2 April 2019
The Scientists’ Choice Award for Life Sciences Video of the Year was awarded to Dr. Adam P. Rosebrock, Assistant Professor of Pathology and Member, Cancer Center, Stony Brook School of Medicine. The winning video, first published on The Scientists’ Channel, details Rosebrock’s work towards developing new assays using the latest mass spectrometry technology from Agilent Technologies to advance the field of small molecule metabolomics.
I'm Adam Rosebrock, an assistant professor at Stony Brook University in the Department of Pathology and also a member of the University Cancer Center. In addition to my own research, I'm tasked with setting up small molecule mass spectrometry for the greater university and cancer center community. That includes outreach such as teaching here at Cold Spring Harbor where I'm guiding the next generation of small molecule mass spectrometrists and teaching them all about gas and liquid phase chromatography and mass spectrometry.
So why is metabolomics an exciting field to work in? For me, it's because we now have the tools that let us ask questions that had been existing for a long time in the community. I'm amazed by how much we don't know about metabolism. Every time we perform an experiment we have the chance of opening up cells and finding new compounds we didn't expect to see there or finding entirely new reactions that no one had thought to map previously.
With the tools for untargeted metabolomics, in particular, we can now measure new compounds and characterize entirely new pathways. What are the challenges in metabolomics research at present? I think the single biggest problem is the lack of association between genome and small molecule contents and reaction rates inside cells, this is a problem I don't think we can actually ever fix computationally.
We can sequence genomes quite easily at this point, we can predict the transcripts and the proteins that genome can make or missing that next step which is predicting the small molecule contents and the biochemical reactions happening inside cells of a given genome sequence. That's a great opportunity for small molecule metabolomics because we have a great business for the foreseeable future in actually measuring what's happening inside cells using mass spectrometry.
I think there are also some very focal challenges in metabolomics at present, the first is separation chemistry. The other challenge that my group really sees as fundamental is one of data analysis. Our workflow is really based around addressing the challenges I just described in mass spec metabolomics. The first thing that I do is really sit down with every student or collaborator that wants to use small molecule mass spectrometry and determine if mass spec metabolomics is the right technique for them.
Once we decided that mass spec metabolomics is the right tool, I'll sit down and establish if we have the right chromatography to separate the compounds of interest. When possible, we like to start off with our off-the-shelf chromatographies to have some standardized method we can use and especially connect to other instruments being performed in my lab and in other labs. If we decide that new methods have to be developed, there's a whole step of chromatographic separation, optimization, maybe building entirely new separations.
In collaboration with Agilent Technologies, my group has developed a chromatographic separation for central carbon metabolites that lets users very quickly move from having a new instrument, having an instrument that's actually generating data useful for their scientific questions. These standardized or commoditized methods also enable users to directly compare their results with other people in the field and now we can start generating data between multiple sites.
My lab uses a combination of liquid chromatography and gas chromatography, chemical separations, we couple that to unit mass instruments like GC mass spectrometer. Our favorite place to be is with accurate mass full scan instruments, primarily time of flight and quadruple time of flight machines. These TOF and QTOF instruments let us go back and mine data we didn't think to look for in the first place, have phenomenal sensitivity and are in general are Jack of all trades for the lab.
For applications, we're looking for very small amounts of compounds or looking just for one or two analytes in a sample. We'll use a triple quadruple instrument for very exquisite targeted, specific and selective quantitation. Once we've collected data, the fun really begins. At that point, we do initial quality control with vendor software. These are programs that are loaded onto the computer at the time your mass spectrometer is checked out but very quickly for our large experiments we've moved to in-house software that lets us extract data from raw mass spectral files across tens or thousands of samples and then eventually pull that into further statistical analysis using software like the AR software suite or other programs possibly even Excel once you're down to fully distilled mass spectral data.
I think it's a great time to be in this business. We're now at a place where we've moved from development as a primary goal into actually using metabolomics to address biological questions. In my lab, we're now for the first time, able to address questions spending hundreds or thousands of samples quantitatively using our internal reference, using our custom-built software and using the increasingly robust mass spectrometers and chromatographic separation that are becoming available every year from vendors.
In my group, we're continuing to build new chromatography, we have new compounds that we'd like to better separate, we have more complex samples coming in we'd like to figure out more compounds from. And so we continue to build new chromatographies to expand existing separations and also add entirely new kinds of chemistry to what we can measure.