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The relative strengths and weaknesses associated with different mass spectrometer configurations can make selecting the right instrument a challenging endeavor for a prospective buyer. This article will discuss the basic components of mass spectrometers, integral specifications, and key considerations necessary for selecting the right mass spectrometer to suit the buyer’s clinical needs.
Mass spectrometry measures the mass-to-charge ratio (m/z) of an ion. The mass-to-charge ratio is useful in identifying compounds because the molecular or atomic weight of each molecule or atom is unique and intrinsic to the sample introduced. The signal measured by the mass spectrometer is the ion current, or the flow of ions filtered by mass, received by the ion counting or A-to-D conversion detection element.1
Quantitative mass spectrometry
Ion currents measured by the mass spectrometer can be made proportional to the amount of sample introduced to the instrument. Thus, quantitative measurement is made possible. In a clinical application, quantitative mass spectrometry strategies are central to identifying biomarkers indicative of disease. Typically, quantification is achieved by measuring the ion current response of an internal standard relative to that of the test sample. Whether the mass spectrometer analysis requires quantitative or qualitative measurement, every sample analyzed should contain a known amount of internal standard to be used as a reference for determining the sensitivity of the assay.
Ionization techniques
Regardless of the vendor or type, all mass spectrometers share the same set of processes. The test sample must be converted to an ionized gas. Depending on the native state of the sample, different ionization techniques can be used. Samples in gas or liquid phase can be ionized by gas chromatography or liquid chromatography, respectively. Samples introduced in a solid state can be converted to ionized gas using matrix-assisted laser desorption ionization (MALDI). MALDI uses a coherent laser to excite, vaporize, and ionize the solid-phase sample from a crystalline matrix. The efficiency requirements, number, distribution, and type of ion created will vary depending on the ionization method (Table 1).
The analyte of interest and internal standard should ideally be equally influenced by ion source events and the ratio of their ion currents unchanged. For this reason, the physicochemical properties of the target analyte and internal standard should be as close as possible. Naturally, the ideal choice for internal standard is a stable, isotype-labeled form of the compound of interest, with a distinguishable mass, also known as an isotopomer.
Table 1 - Ionization and mass separation methodologies
Mass analysis
Once the sample has been converted to gas-phase ions, including those indicative of the target analyte and those not of interest, they travel to the mass analyzer. Here, different ions are separated based on their mass-to-charge ratio. There is significant variation in the separation mechanism of ions (Table 1b), the details and capabilities of which cannot be fully explored within the scope of this article. The operational parameters of mass spectrometry are conserved, such as mass scale calibration, resolution, limit of detection, limit of quantification, analytical range, precision, and ease of optimization. Important mass spectrometer specifications are given in the Mass Spectrometer Features Checklist.
Signal detection and analysis
Once filtered by the mass analyzer, ions bombard the ion counting or A-to-D conversion detection element. The detector gives an electronic response proportional to the number of targeted ions present. This electronic signal is then interpreted by a computer with bioinformatic software, which controls various instrument parameters and processes the data.
Key purchasing considerations: Defining requirements
Throughput
There are many factors that play into the number of tests that can be performed in a given time period, making overall mass spectrometry throughput difficult to quantify. “Throughput encompasses several system performance factors, analytical speed, system sensitivity, system robustness, tune stability and overall reliability,” explained Meredith Conoley, GC & GC-MS Marketing Director at Bruker Chemical and Applied Markets Division (Fremont, CA). “Placing too much emphasis on any one criterion, such as speed, can lead to a choice that does not deliver in another key area such as robustness. Any performance gains due to speed can be lost due to time spent with frequent maintenance (cleaning) or re-tuning. This results in loss of overall sample throughput.”
Sensitivity and selectivity
The sensitivity of a mass spectrometer is defined as the change in response of the measuring system divided by the corresponding change in stimulus. The selectivity of the instrument is defined as the ability to accurately measure the target analyte in the presence of interferences.1 “Sensitivity is a very important parameter for clinical assays,” continued Conoley. “The complexity of the sample matrix requires an equal evaluation of system performance based on selectivity. Sample complexity and required detection limits can be the determining factors on whether one chooses a GC-MS single quadrupole system or GC-MS triple quadrupole [TQ]. While GC-MS TQ may be a more expensive investment, the inherent increase in selectivity provided by MRM [multiple reaction monitoring] capability of the triple quad can increase the ability to detect target analytes to the sub-ppb range in the complex sample matrix in clinical applications. Selectivity is as important a factor in system performance as sensitivity for the detection of analytes in complex matrices.”
Precision and accuracy
Precision and accuracy vary between mass spectrometer technologies and vendors. Measurement precisions of 1% can be achieved using quadrupole mass spectrometers, while precisions of better than 0.1 ppm will require high-resolution instruments.
“One of the major challenges is that different workflows create different kinds of information, making reproducibility and standardization a key to confidence in generated results,” Conoley said. “Standardized analysis methods and validation techniques in SOPs significantly assist in comparability of results.”
Data processing software
Mass spectrometer software controls the hardware, methods of analysis, data analysis, and result storage. “All systems will process the data they acquire but not all systems are designed with the workflow of the laboratory in mind,” Conoley pointed out. “Does it support critical compliance requirements; does it support connection to my LIMS? These are key issues to investigate beyond the basics of data processing.”
Other considerations to take into account prior to purchasing a mass spectrometer are size, ease of use, instrument versatility, and quantitative (diagnostic) versus qualitative (screening) analysis needs.
Validation
Once the buyer’s needs are well understood, the stakeholder should clearly define the target analytes and develop or obtain a test compound. Based on the target analytes and the defined requirements, the purchaser should identify a mass spectrometer and the necessary components. It is critical to match compatible ionization modes, resolution required, and the ionization method. Using the test compound, a demonstration session with vendors of interest should take place at which the vendor tests key metabolites and evaluate sensitivity, precision, ease of optimization, and data processing software. Ideally, this demonstration should be paired with a parallel analysis using a validated method. Additionally, interinstrument comparisons should be performed on identical samples under identical conditions. The evaluation should be on site if possible. Lastly, instrument software, data processing, and result interpretation should be thoroughly demonstrated and explained by the vendor.
Important price points
Conoley suggested that buyers not be swayed by the initial cost of the instrument. “Evaluate in terms of what additional technical capabilities it may bring to the workflow in terms of cost per sample, increased productivity, etc.,” he said. “In terms of GC-MS, investing in a more expensive GC-MS TQ system may actually allow an overall reduction in cost per sample due to reduced sample prep costs, reduced sample collection costs, etc., due to the improved quantification capability of MRM over SIM [secondary ion mass] or full scan in a GC-MS SQ [single quadrupole].” References from laboratories that own the current model can also be helpful when trying to understand the workload impact, reliability, and ease of use of an instrument already in use.
Conclusion
When purchasing a mass spectrometer, certain regulatory issues (i.e., certifications and procedures) may drive selection of devices used in the clinical environment. “Users must be aware of these requirements and ensure the equipment they are purchasing has met these requirements,” Conoley said. It is also important to consider the reputation of the vendor itself. Conoley points out that non-product issues must be evaluated in addition to technical needs. These include the industry knowledge and depth of experience of the company.
Reference
- Mass Spectrometry in the Clinical Laboratory: General Principles and Guidance; Approved Guideline. CLSI document C50-A; Wayne, PA: Clinical and Laboratory Standards Institute, 2007.
T. Keith Brock, BS, is a Contributing Writer, American Laboratory/Labcompare; e-mail: [email protected] . The author would like to thank Meredith Conoley, GC & GC-MS Marketing Director, Bruker Chemical & Applied Markets Division (Fremont, CA), for his contributions to this article.