Quantitative Biological and Clinical Mass Spectrometry

An Introduction

Anthony I. Mallet

• Publisher: Wiley
• Year: 2018
• Print ISBN:
  9781119281207
• eBook ISBN:
  9781119281214
• Edition: 1
• Topic: Physical Sciences
• Subtopic: Spectroscopy & Spectrum Analysis

A basic overview of mass spectrometry relevant to life and health science applications, illustrated throughout with relevant case studies

This introductory text provides information and assistance to new users of mass spectrometry (MS) working in clinical or biochemical fields who are faced with implementing and designing quantitative mass spectrometric assays for a variety of classes of molecules of biological interest. It presents a detailed discussion on how to optimize measurement parameters for a candidate reference quantitative analysis, including calibration procedures, sensitivity, reproducibility, speed of assay and compliance with regulatory authorities. 

Quantitative Biological and Clinical Mass Spectrometry uses examples where development has not been immediately successful but where unforeseen problems have arisen and describes the strategies used to solve these. Advances in addressing the very large numbers of clinical samples that arise on routine screening programs such as those involved in inborn errors of metabolism studies are discussed. Direct mass spectrometric based analyses applicable to point of care testing (POCT) situations are also covered. The book concludes with a short section on possible novel developments, bibliography, references, and a glossary of terms.

• Shows how the presence of false results can be detected and understood
• Describes the 'parts' of modern instruments from sample introduction through ionization, mass analysis and detection, and the variety of techniques of tandem mass spectrometry
• Discusses the requirement for specificity in an assay method
• Fully illustrated throughout
• Highly relevant to all key areas of mass spectrometric analysis

Quantitative Biological and Clinical Mass Spectrometry appeals to those newly exposed to the use of combined chromatography and mass spectrometry for analysis of biological material and to scientists experienced in automated clinical analysis using immunoassays or who are new to mass spectrometry.

1
The Instrument: Ion Creation

1.1 Introduction

The modern mass spectrometer is made up of a number of distinct units. Figure 1.1 shows the disposition of these.

Figure 1.1 Schematic of a modern chromatography/MS instrument.

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Modern instruments are more closely integrated in the sample introduction and mass analysis sectors. The whole system should, ideally, be controlled by a single data system.

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1. A sample introduction device.
2. An ionising source. The original source was a vacuum in which a beam of electrons impinging on the analyte molecules induces ionisation. It is only able to accommodate gaseous samples but is still popular as the source of choice to link with gas chromatography (GC). Today, ion sources operating at atmospheric pressure are equally popular and are easier to interface to samples in condensed phases and with liquid chromatography.
3. An analyser in which to separate ions differing in mass and to transport them to the detector. A range of different configurations is in use; those that are best designed to produce quantitative analysis results are described here. Multiple analysers are used for MS/MS.
4. A detector. This records the impact of the ions from the analyser(s) and stores the signals.
5. A dedicated data system to operate the instrument and collect and collate the data.

The data output from the instrument can take two forms: one will be a so‐called mass spectrum, which is a plot of the ions detected against their intensities, and the other is a chromatogram in which the intensity of one or more preselected ions is plotted against the elution time from the sample introduction system.

The whole operation of a mass spectrometer is based on the production of charged molecular species, ions, in which the electrical charge acts as a ‘handle’ by which the ion can be moved through electrical fields. All measurements are made on the ratio of the mass‐to‐charge of the ions. It is essential, in order to prevent losses of ions through mutual charge repulsion and collision, that they are retained in a vacuum, and all mass spectrometers will have efficient vacuum pumps attached. A fundamental problem with all mass spectrometers will be the inherent tendency of bundles of ions all carrying the same charge to repel each other; much progress has been made in clever ion focusing systems to enhance their transmission through the parts of the instrument.

1.2 Sample handling

The process of ionisation is often made simple and efficient if the sample can be produced as a uniform homogeneous purified product. A mixture of too many different species can inhibit the formation of the expected ions, a process referred to as matrix inhibition or interference. This will be treated in detail later. In early work it was, and still is, simple to link the eluent from a gas chromatograph column straight into the ionisation chamber and then to perform a direct analysis of the material in real time. Chapter 3 discusses those methods most applicable to quantitative analysis.

It was only in the 1980 s that the same process became feasible for liquid samples, especially solutions, such as occur in the eluent of a high‐performance liquid chromatography (HPLC) apparatus. In an integrated mass spectrometer, an important part is the GC and or HPLC equipment. It should be controlled by the same data system as the rest of the instrument.

Later on we discuss a number of recent developments whereby the sample can be accessed directly, in the open laboratory, ionised and desorbed into a vapour.

1.3 Vacuum ion sources

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The need to provide reliable and precise vacuum conditions is vital for the operation of...

Table of contents

• Cover
• Introduction
• 1 The Instrument: Ion Creation
• 2 The Instrument
• 3 The Mass Spectrum
• 4 Sample Handling Prior to Ionisation
• 5 Establishing Optimum Specificity
• 6 Quantitative Analysis with Mass Spectrometry
• 7 Examples of Quantitative Analysis
• 8 Rapid Clinical Analysis
• A Simple Mass Spectrometry Fragmentation Mechanisms
• B Some Simple Derivatisation Methods
• C Acronyms and Glossary of Common Terms
• D Simple Statistics
• E Helpful Web Links
• Bibliography
• Index
• End User License Agreement