Wavelength Tech Forum: Mass Spectrometry

May 30, 2007

This month's Technology Forum looks at the topic of Mass Spectrometry and the trends and issues surrounding it. Joining us for this discussion are Chad Ostrander, with Hitachi High Technologies America, Dale H. Patterson, with Applied Biosystems, and Lester Taylor, with Thermo Fisher Scientific.

This month's Technology Forum looks at the topic of Mass Spectrometry and the trends and issues surrounding it. Joining us for this discussion are Chad Ostrander, with Hitachi High Technologies America, Dale H. Patterson, with Applied Biosystems, and Lester Taylor, with Thermo Fisher Scientific.

How has tandem mass spectrometry changed the MS field?

(Ostrander) Tandem MS in both time and space has dramatically changed the productivity of MS related research. Throughput and molecular information are critical; tandem MS provides both. Molecular structure studies, traditionally limited to complex NMR systems, are now routinely performed on relatively inexpensive online Tandem LC-MS platforms.In the field of proteomics specifically, Tandem MS techniques have contributed to the recent transformation from general protein identification applications to the more specific targeted proteomics experiments. The end result is more “real” applications for drug discovery and biomedical research.

(Patterson) From a quantitative standpoint, tandem mass spectrometry is absolutely fundamental to deliver the specificity and sensitivity for assays required in so many markets including pharmaceutical DMPK, clinical research, forensics, food, and environmental analysis. The use of tandem MS enables researchers to create safe and effective drugs that improve the human condition, keep food supplies safe, shed light on sports doping and protect the environment, just to name a few.Qualitatively, it allows researchers to identify potential toxic metabolites that our bodies form from the drugs that we take as well as to identify the proteins that are key players in mechanisms of disease.

(Taylor) Over the last 30 years there have been significant changes in the use of tandem MS. Originally developed for high end MS research in areas such as the study of metastable ions and ion kinetic energy spectra, tandem MS instruments have become standard equipment in a wide range of labs including those carrying out proteomics research, metabolite identification, and drug bioanalysis. Now highly integrated and automated, today's tandem MS instruments are capable of 24/7 operation. Notably the triple quadrupole tandem MS instrument is the preferred system for quantitation in drug discovery and development as well as monitoring contaminants in food and environmental samples.

What effect has isotope dilution mass spectrometry (IDMS) had on the MS field?

(Patterson) Isotopically labeled standards allow us to account for issues such as ionization suppression, chromatography variability and matrix effects, thereby enabling the required precision in quantitative analysis by mass spectrometry, not afforded by surrogate internal standards. This, coupled with the state-of-the-art tandem mass spectrometers on the market today, yields unparalleled confidence in the results for all the quantitative application areas I previously mentioned. One interesting trend that we are seeing expands the field of IDMS to protein-based biomarker discovery. Some new reagents, for example, use heavy isotopes in a unique way giving the researcher the capability to find proteins that are quantitatively up- or down-regulated in a disease or perhaps ones that are indicative of a toxicological effect of administering a drug.

(Taylor) It has allowed the development of rigorous absolute quantitation methods for target compounds in several fields of application: drug discovery and development, food and environmental analysis, and clinical and toxicology screening.

What further effect do you see MS having on the biomedical industry?

(Ostrander) Investments in biomarker research and targeted proteomics will continue to grow. Today’s mass spectrometers are faster, cheaper, and more powerful than ever affording an immense amount of real-time data. MS will continue to be a primary tool for biomarker research, but the most growth in the market will be due to the resulting clinical applications.High speed TOF based MS analyzers with UHPLC chromatography systems will have a significant presence in new clinical MS methods for biomarker screening. The current hardware capabilities combined with the increasing number of known biomarkers will greatly enhance clinical analyses with regards to throughput, analysis time, and the number of components/diseases tested.

(Patterson) Mass spectrometry is playing an increasing role shortening the time needed to bring drugs to the market where they can help people live longer and more fruitful lives. For example, we are seeing technologies emerge, like a MALDI interface on a triple quadropole, that will help speed up the quantitative analysis for pre-clinical early ADME labs more than 25 fold from what is available today. Biomarkers have a real, yet unrealized, potential in this field. The use of tagging reagents is enabling scientists to find putative markers of disease and toxicology through discovery experiments. From there, we are seeing the application of traditional small molecule quantification instruments to verify the real utility of the marker in larger cohorts. This is a really exciting time to be in the mass spectrometry field and watch these enabling tools be used in such productive ways.

(Taylor) Continued use in the areas of drug discovery and development. Now that biomarker research is more widespread MS is the instrument of choice for (1) biomarker discovery, (2) method validation, and (3) target biomarker quantitation. Ultimately MS technology may become more routine in clinical monitoring.

What improvements could be made to mass spectrometers to make them more efficient?

(Ostrander) Of course, fundamental improvements in resolution, mass accuracy, mass range, sensitivity, and scan time are always beneficial to some applications. In general, however, it has become clear that two areas offer the most room for improvement: automation and quantitative analysis.Automation is the key to true high throughput research and applications. To date, the high-end MS instrumentation typically used for biomolecule research has limited automation capabilities. Automation will significantly increase the amount of information generated.While there have been recent improvements in the quantitative capabilities of MS instruments, they continue to be relatively limited in terms of dynamic range, accuracy, and reproducibility. This is particularly critical when dealing with proteins that can exist in situ at concentration ranges of 10+ orders of magnitude.

(Patterson) If we look at the area of sensitivity, we are seeing a plateau in what we can do from an ion creation and transmission standpoint. For example, in triple quadropoles, we have seen a 6 order of magnitude increase in sensitivity over the past 25 years which has been driven largely through improvements in these two areas and are nearing the physical limits. I think the real immediate wins here are in how we better use the tools that we have. We can make improvements in the area of more intelligently selecting and fragmenting parent ions. For example, we can do a better job of filtering background ions, selecting ions in MetID experiments that have the same mass defect, dynamically adjusting the collision energy to assure quality MS/MS spectra and exiting out of MS/MS acquisition when the data quality is high enough. Orders of magnitude improvements in metabolite and protein ID experiment have been demonstrated simply by applying these types of software tools to the instruments that we have today.Another area is in the unique coupling of mass analyzers to take advantage of their inherent strengths. For example, we are seeing researchers starting to take full advantage of the coupling of a triple quadropole with a linear ion trap to couple a highly sensitive and selective triple quad scan like an MRM with a highly sensitive full scan MS/MS of the linear ion trap to give the researcher qualitative ID and quantitative precision and accuracy simultaneously.

(Taylor) Ongoing developments for greater sensitivity combined with more selectivity. More automated and routine operation for high throughput sample analysis. Intelligent data processing and automated reporting functions.

What other significant developments of interest have happened in the MS field recently?

(Ostrander) Improvements in MS hardware and application are ongoing. However, a fairly recent development that has drastically changed the overall performance and capability of MS is related to the instrument scan speed. High speed analyses are no longer limited to simple low sensitivity MS techniques such as TOF or SIM/SRM scanning techniques. Now, high sensitivity full mass range MS/MS analyses can be performed on virtually all high-end LC-MS platforms. Reduced scan times, on the time scale of UHPLC separations, allow for comprehensive, online interrogation of samples using powerful gas-phase fragmentation methods such as CID and ECD.

(Patterson) I can point to a number of key developments in the areas of instrumentation, workflows, and software that have been enabling.On the instrument side, the introduction of the first MALDI ionization source on a triple quadropole mass spectrometer is designed to be a real quantum leap in speed for quantitative analysis of small molecules. This new platform is intended to improve the speed of analysis by 25 fold allowing researchers the ability to screen more drug leads through a standard set of assays in the early ADME pre-clinical stage.The biomarker area has been aided by new workflows enabling precise quantification to verify putative markers. Researchers are using hybrid triple quadropole linear ion traps to develop and then implement highly sensitive methods to screen larger sets of samples to determine if the potential biomarker is ready to take to a validation stage. Until now, this has been a gap in the biomarker pipeline. One of the most important areas of improvement has been in software. The power of quantitative tandem mass spectrometry is finally reaching the “black-box” stage, thanks to easy-to-use software interfaces that enable technicians to use pre-canned methods to monitor the safety of our water and food supplies and do routine drug testing.

(Taylor) Routine use of accurate mass and high resolution has lead to a significant increase in the confidence with which MS is used for several applications, whether it is peptide identification, metabolite identification. More recently the introduction of ETD has significantly change proteomics. It is a powerful new tool for peptide structure analysis and protein identification, including post translational medications, and top and middle down sequencing. The introduction of FAIMS technology (high Field Asymmetric waveform Ion Mobility Spectrometry) on ion traps brings new degrees of separation to mass spectrometry. This allows users to select ions based not only on mass, but also based on charge state, molecular shape, and conformation.

What do you think?

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