MALDI Time-of-Flight Mass Spectrometry

Introduction

MALDI is a soft ionization technique used in mass spectrometry (MS) that produces rapid and efficient ionization of a wide variety of molecules (see Figure 1). MALDI uses a laser energy absorbing matrix to produce ions from molecules having molecular weights ranging from 100's to 1000's of Daltons with minimal fragmentation. It has found increasing application over the past 30 years, especially for the analysis of biomolecules, e.g. proteins, peptides, DNA, and polysaccharides, and other large organic molecules, e.g. polymers, dendrimers and other macromolecules. These molecules tend to be fragile, undergoing excessive fragmentation when ionized by more conventional methods.

MALDI is similar in character to ElectroSpray Ionization (ESI) in that both techniques produce low molecular fragmentation on ionization. A primary difference between MALDI and ESI is that the former typically produces ions with a net single charge and this enables simple determination of molecular mass for most compounds. However, it can also limit the ability to analyze the largest macromolecular proteins. MALDI ionization requires three steps:

  • A sample is mixed with a suitable matrix material and applied to a metal plate. As the mixture dries, crystals are formed on the sample material, that are critical for efficient ionization.
  • A pulsed laser beam impinges on the sample, causing desorption of the sample and matrix material. The matrix material breaks down in this process to produce gas phase ionic species.
  • Finally, the analyte molecules are ionized via protonation or de-protonation in the hot plume of ablated gases. The ions are then accelerated into a mass spectrometer system for mass analysis.
The MALDI process
Figure 1. The MALDI process (Figure used with permission of Dr. Paul Gates, University of Bristol UK).
While MALDI units are most commonly coupled with Time-of-Flight (ToF) mass spectrometers (Figure 2), in recent years they have been associated with a wider variety of instrument types that enable their use in both advanced research, medical and clinical applications. MALDI units have been used with:
  • Laser technologies
    • UV gas lasers, e.g., nitrogen laser operating at 337 nm
    • DPSS Nd:YLF or Nd:YAG lasers with frequency-tripling (operating at 349 nm and 355 nm, respectively)
  • ToF MS
    • Axial ToF . affordable performance, speed, ease-of-use
    • Orthogonal-acceleration ToF . higher performance, specificity, flexibility
    • Ion mobility spectrometry orthogonal-acceleration ToF - highest specificity
  • Tandem quadrupole MS
    • Industry standard quantitative platform (despite limited MALDI linearity)
MALDI MS analysis, with an axial ToF detector
Figure 2. MALDI MS analysis, with an axial ToF detector.

Typical applications for MALDI MS include, but are not limited to:

  • Biochemistry and molecular biology, e.g., proteomics, glycomics
  • Organic chemistry
  • Polymer analysis
  • Microbiology
  • Medical and diagnostics

Impact of Photonics on MALDI

The incorporation of photonics technologies into MALDI MS instruments dramatically improves the sensitivity, throughput, and ease of use of these systems. This has led to their increased deployment in biological and polymer research where MALDI systems have provided an attractive alternative to older, more complicated high-performance Liquid Chromatography (LC/MS) units. Additionally, improvements in laser technology have enabled high-speed analysis, which significantly increases the throughput of protein identification in proteomics research laboratories. More recently, as ESI-LC/MS has become the preferred method for proteomics, MALDI MS has found wider application in microbiological, pharmaceutical development and medical-related applications. The key advances in laser technologies and their consequences for MALDI methods are:

  • Tunable wavelengths - more efficient ionization of biomolecules
  • Increased repetition rates - speed of analysis
  • Improved lifetime and cost of ownership - reduced complexity and increased reliability

MALDI MS units have been increasingly used in imaging applications where the highly localized nature of laser ablation allows the determination of the concentrations and spatial locations of compounds of interest in a wide range of sample types, especially biological tissues. The key photonics technological drivers for advanced imaging of compounds in tissues include:

  • Laser repetition rate (> 1 kHz) - this enables fast processing of large tissue/sample sections
  • Laser energy profile - this ensures minimal denaturation of a sample below the ionization threshold which, in turn, improves sensitivity, particularly for spot sizes below 20 µm
  • High precision optics and motion control - this provides the ability to reliably measure spatial locations under 10 µm

Imaging applications are currently driving the equipment requirements in research environments within the life science and clinical communities. The critical equipment characteristics for MALDI units in these applications include:

  • High resolution imaging - sub-20 µm spatial resolution
  • Sample stage control - fine X/Y control, high robustness and reproducibility
  • Speed of acquisition - using small spot size, GHz pulse rate, top-hat energy profile
  • Flexibility - coupling of lasers by optical fiber

Explorer® One™ 349 nm lasers have been specially designed with MALDI customers in mind. With the Explorer One series, users can rely on reliable ionization process and eliminate unwanted side effects, thus increasing the longevity of the laser and avoiding unnecessary service interruptions. Explorer One lasers are offered at a variety of pulse energy values from 30 to120 mJ and typical pulse width of < 5ns.


MKS Semiconductor Handbook Cover

For additional insights into photonics topics like this, download our free MKS Instruments Handbook: Principles & Applications in Photonics Technologies

Request a Handbook