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Mass Spectrometry in Biological Research – A Guide for Beginners

Image shows a laboratory with mass spectrometry facility to represent using mass spectrometry in biological research

Mass spectrometry, also referred to as mass spec or MS, is an analytical technique that is becoming increasingly important in bioscience research. This article will introduce you to mass spectrometry in biological research, explain how it works, and how it could be useful in your research.

What Is Mass Spectrometry?

In a nutshell, mass spectrometry accurately measures the mass of different molecules within a sample. Even large biomolecules like proteins are identifiable by mass, which means that biologists can perform some very interesting experiments using mass spectrometry, potentially adding a new dimension to their research.[1]

Mass Spectrometry in Biological Research – is it Useful?

Yes, if you want to:

  • Identify molecules in a mixture;
  • Detect impurities in a sample;
  • Analyze a purified protein;
  • Study the protein content of a sample of cells.

How Does Mass Spectrometry Work?

Before you get started using mass spectrometry in biological research, you should make sure you understand the basics of how the technique works. There are three key stages to a mass spectrometer, and we discuss each in more detail below.

1.  Ionization

Molecules in a sample are vaporized (converted to the gas phase by heating). Then, an electron beam bombards the vapors, which converts the vapors to ions. Because mass spectrometry measures the mass of charged particles, only ions will be detected, and neutral molecules will not be seen.

Ions are created by giving electrons to a molecule (producing a negatively charged ion) or taking electrons away from a molecule (producing a positively charged ion).

Note: A sample can only be analyzed by mass spectrometry if it can be vaporized without decomposing.

2.  Acceleration and Deflection

Next, the ions are sorted according to mass in two stages – acceleration and deflection.

Acceleration is simply attraction. The positive ions created in the ionization stage accelerate towards negative plates at a speed dependent on their mass. In other words, lighter molecules move quicker than heavier ones.

Deflection is the magnetic bit. Here, the ions are deflected by a magnetic field, and the extent of deflection is again dependent on mass. So, ions of different masses travel through the spectrometer at different speeds.

3.  Detection

Ions of increasing mass eventually reach the detector one after another, and then it’s over to the computer to provide a spectrum. The output will look something like in Figure 1.

Mass Spectrometry in Biological Research – A Guide for Beginners

Figure 1. A simplified spectrum of pentane (CH3CH2CH2CH2CH3) produced by a mass spectrometer.

Note: This simplified mass spectrum has been drawn using data from the Spectral Data Base System for Organic Compounds (SDBS) at the National Institute of Materials and Chemical Research in Japan.

Omission of minor lines with peak heights of 2 % or less of the base peak (the tallest peak) provides clarity.

Reading The Spectrum

First, you should note that the x-axis denotes not just ‘mass’ but ‘m/z’, where m is the mass and z is the charge. What the spectrometer actually measures is the mass/charge (m/z) ratio. Sounds complicated, but it’s not – because since the majority of ions produced during mass spectrometry have a charge of 1, m/z is normally equal to the mass of the molecule. Simple!

Is Mass Spectrometry in Biological Research Worth It?

In a word, yes! Mass spectrometry provides accurate weight measurements for your bio- (or other) molecules, which can be used to:

  • Give a good estimate of the purity of the sample (i.e. whether there are one or more molecular species in your sample and what ratio those species are in)
  • Monitor reactions, sequence amino acids, and oligonucleotides as well as give information on protein structure
  • Identify and quantify samples

Mass spectrometry has enabled biology to move from identifying single proteins to proteome-wide characterization and quantification. With the development of this tool, researchers can now feasibly begin defining biochemical pathways on a kinetic basis, which will help us understand the mechanistic response of cells to changing environments.

 Practical Uses of Mass Spectrometry in Biological Research

Drug companies and researchers are using the technique for drug discovery, for gaining information on drug metabolism, and for pharmacokinetic studies.[2] The study of proteins through mass spectrometry is now aided by information obtained from genome sequencing projects.[3] The technique has also found medical uses, such as neonatal screening, blood analysis, and drug testing. These uses also make mass spectrometry (when coupled with other analytical techniques) a powerful tool in forensic analysis.

How Do I Go About Using Mass Spectrometry in Biological Research?

If you feel that mass spectrometry is something that could be of use in your research, check whether your department or college has mass spectrometers available for use. This might be either as a ‘walk-up’ service where you book a slot and run your own samples or as a technician-run service. Training will always be provided for these services so ask around.

While the number of uses is large, the instrument itself is not – normally the machine can fit on a benchtop.

But Remember This Before You Use a Communal Mass Spectrometer

The most important factor in getting accurate results is keeping it clean and free of contaminants. As many a chemist will testify, this is often an issue with communal mass spectrometers – especially if the user before you did not clean the column properly. It’s wise to bring your own solvent to clean the column before (and after!) use.

On the other hand, also note that accidental contamination during sample preparation can mess up your results, and keeping a tab of these mass spec contaminants can help your experiment.

Want to Know More?

This article was a brief introduction to the basics of mass spectrometry, but it doesn’t end here! There are various ways of producing the required ions, and the method chosen depends on the nature of the sample molecule. You can find out more information in the references below. Try out the technique and let us know what new applications it finds in your research.

References

  1. Baker M. (2010) Mass spectrometry for biologistsNature Methods. 7:157.
  2. Ashcroft AE. An Introduction to Mass Spectrometry.
  3. Yates JR. Mass spectrometry in biology. In: Encyclopedia of Life Sciences. pp. London: Nature Publishing Co.; 2001. 1–5.
Originally published February 16, 2011. Reviewed and republished June 2021.

15 Comments

  1. callan on October 3, 2018 at 1:57 pm

    k



  2. Glaxo Smith Kline on April 17, 2018 at 7:18 pm

    Great! thanks mate



    • Uwase on November 8, 2019 at 7:22 am

      It is good the way you have explained but be on ionisation we need more details thanks.????



  3. Jenny on January 20, 2018 at 5:47 pm

    Thanks a lot for all your helpfull articles! Here, I was a little confused by your use of mass spectrometry and mass spectroscopy. Is there another article (which I missed) that explained the difference? Because there is a difference, but in this article it sounds like it’s the same…



  4. Chris on November 3, 2017 at 5:11 pm

    This is an excellent article right in line with the spirit of BitesizeBio.

    There is however a significant grammatical error quoted here “Because mass spectroscopy measures the mass of charged particles, neutral molecules only ions will be detection, and neutral molecules will not be seen.”



  5. Matteo on November 10, 2016 at 1:34 pm

    Hello, This article has really been helpful to me for a school project!!
    One thing I do not entirely grasp is how you can tell which molecules are being separated, and which molecules are which at the end. For example this machine can also be used to test athletes to see if they have been taking drugs. How does it do that?
    Thank you again for all this information



  6. Jasper Whiteside on October 27, 2016 at 5:41 pm

    I’m glad the article mentions that a mass spectrometer can be used in forensics because I was trying to remember where I had heard of it before. The answer is from a TV show that I used to watch. It really is fascinating how powerful these machines are in analyzing biological materials.



  7. Allison Ross on February 28, 2011 at 1:53 pm

    Lawrence – that great to know! We’ll be taking the series through the basics first, looking at why this particular technique works in the way that it does. Parts 2 and 3 are coming up shortly which explain some of the different methods of ionizing your sample and compare ESI with MALDI. Later, we’ll be looking at more specific uses and functions in a biological frame. Keep telling us what you want, and we’ll do our best to find the information for you. Thanks!



    • incommunicada on July 21, 2012 at 8:09 am

      Do you have an article on how to read ESI/MALDI spectra? I haven’t fully grasped how yet.



  8. Lawrence Forsberg on February 26, 2011 at 6:13 am

    Tally me as another person who’s really enjoyed the articles lately.

    I’m personally interested in the identification of metabolites in cells under different conditions (drug treatments, knockouts of different genes, etc.) to identify the role of uncharacterized enzymes, and things like post-translational modification mapping.

    Any chance your next article (or another some time soon) will cover any of these?

    Thanks again for the good read!



  9. Carl Saxton on February 23, 2011 at 5:12 pm

    Thanks for your comment. I am glad you enjoyed reading the article. I’d be interested in knowing what part of the article interested you most? Is there any specific Mass spectrometry topic you would like us to cover?



    • veena on October 19, 2012 at 12:41 am

      Hello Mr.Saxton, This article has been extremely helpful to me in understanding how ionization occurs. Thank you very much! However I have various other doubts on how the sample is processed. For instance in HPLC/ESI-MS initially the sample undergoes liquid chromatography. Different compounds gets separated based on their mass and gets eluted at different times based on the retention times. So the elution from the HPLC is discontinuous. Does the MS analyze the sample as and when it gets eluted from HPLC and gives a spectrum for each such event? I hope this question makes sense. I basically want to know what exactly happens to the sample after the ionization process.



  10. Amin on February 22, 2011 at 6:35 pm

    These are really great series. I look forward learning more. Thanks a lot!



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