Liquid Chromatography Tandem Mass Spectrometry (LC-MS/MS)

LC-MS/MS is a powerful analytical technique that is used for nearly all small molecules. With the advantages of high sensitivity and high selectivity, this technology has been widely used in many fields, including pharmaceutical, environmental monitoring, food safety and clinical diagnosis.

Principle of LC-MS/MS

In the LC-MS/MS analysis process, the sample is first separated by LC, and the effluent from the chromatographic column is transferred to the mass spectrometer. The separation by LC is based on the different physical and/or chemical properties of the analytes, such as hydrophobicity, molecular size, differences in functional groups, etc. When the effluent reaches the mass spectrometry ion source, ionization occurs, producing charged particles. These charged particles then migrate under high vacuum through a series of mass analyzers by applying electromagnetic fields. In general, the signal intensity of the ions produced by the mass spectrometer is proportional to the amount of analyte in the sample.

Composition of A Tandem Mass Spectrometer

A tandem mass spectrometer consists of different components. Firstly, an atmospheric pressure ionisation source, typically an ESI source (as shown in Figure 1B) or an atmospheric pressure chemical ionisation (APCI) source (as shown in Figure 1C), coupled by an ion-inlet and focusing component (Q0), which facilitates the transition from atmospheric pressure to vacuum and ion-focusing. Subsequently, a first mass-filtering device (Q1), which leads into a collision chamber (Q2) that can be filled with low-pressure gas for collision-induced dissociation (CID). Next, there is a second mass-filtering device (Q3). Finally, the instrument is equipped with an ion-impact detector (electron multiplier).

Figure 1. Principal components of a tandem mass spectrometer ^[1]. (A) The sample is ionised in the source, passes into the 1st mass filter (Q1), then into the collision cell (Q2), followed by the 2nd mass filter (Q3), and finally the detector. (B) and (C) depict schematically the two principal types of ionisation-sources that are in use in current LC-MS/MS instruments, electrospray ionisation (ESI, B) and atmospheric pressure chemical ionisation (APCI, C).

Multiple Options for Sample Analysis

To enable multiple options for sample analysis, the LC-MS/MS instrument allows the user to perform the following five different experiments within the range of sensitivity and mass resolution of the instrument.

Figure 2. Different modes of operation of mass filtering tandem mass spectrometers ^[1].

A. Full Scan: Scan across the entire (or partial) mass range of two mass filters (Q1 and Q3), while Q2 does not contain any collision gases. The experiment allows the user to see all the ions contained in the sample

B. Product Ion Scan: A specific m/Q is selected in Q1, Q2 is filled with collisional gas so that the selected m/Q produces fragments, and then the entire (or partial) mass range of Q3 is scanned. The experiment allows the user to see all fragment/product ions of the selected precursor ion.

C. Precursor Ion Scan: The entire (or partial) mass range of Q1 is scanned, and the precursor m/z values of specific product ions are selected for monitoring at Q3.

D. Neutral Loss Scan: Scan the entire (or partial) mass range of Q1 as well as the product ion (Q3) to identify all precursors that add product by losing a similar uncharged (neutral) type in all precursors.

E. Selective Reaction Monitoring (SRM): Q1 and Q3 are set to monitor the actual m/z values of the precursor and product ions. Tandem quadrupole mass spectrometry (MS/MS) allows simultaneous monitoring of the precursor-to-product transition for several identical and different analytes.

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Reference

Grebe, S. K. G; Singh, R. J. LC-MS/MS in the clinical laboratory–where to from here? The Clinical biochemist reviews. 2011, 32(1): 5-31.

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