Fast, High Resolution Mass Spectrometry Imaging Using a Medipix Pixelated Detector

In mass spectrometry imaging, spatial resolution is pushed to its limits with the use of ion microscope mass spectrometric imaging systems. An ion microscope magnifies and then projects the original spatial distribution of ions from a sample surface onto a position-sensitive detector, while retaining time-of-flight mass separation capabilities. Here, a new type of position-sensitive detector based on a chevron microchannel plate stack in combination with a 512 × 512 Medipix pixel detector is coupled to an ion microscope.

Advantages of CMOS pixel detectors in mass spectrometry imaging are the ease of data acquisition and data processing (direct image acquisition), the high spatial resolution (possibly enhanced by centroiding algorithms), the single-particle counting mode (noise-free particle counting as opposed to charge integration in CCD-based detectors) and the capability of some detector generations to take time-resolved images (Timepix).

Given these attractive CMOS detector features, we have demonstrated the technical, in-vacuum implementation and capabilities of this novel type of charged-particle detector –a chevron MCP stack in combination with a Medipix2 imaging readout chip- for microscope mode MSI.

High Dynamic Range Bio-Molecular Ion Microscopy with the Timepix Detector

Highly parallel, active pixel detectors enable novel detection capabilities for large bio-molecules in time-of-flight (TOF) based mass spectrometry imaging (MSI). In our detector development project, a 512 × 512 pixel Timepix assembly combined with chevron microchannel plates (MCP) captures time-resolved images of several m/z species in a single measurement. The use of a MCP-Timepix assembly delivers an increased dynamic range of several orders of magnitude. The Timepix returns defined mass spectra already at sub-saturation MCP gains, which prolongs the MCP lifetime and allows the gain to be optimized for image quality. The Timepix peak resolution is only limited by the resolution of the in-pixel measurement clock. Oligomers of the protein ubiquitin were measured up to 78 kDa.

Microscope mode MSI enables fast, high resolution, large area imaging provided that a fast, two-dimensional, time- and position-sensitive detector is used to record high quality molecular images.

In view of the importance that imaging techniques have acquired in biological, bio-molecular and bio-medical MSI, it is remarkable that the basic technology, i.e. 2D image detection and separate arrival time acquisition, has hardly changed over the years. Only recently, the use of in-vacuum pixel detectors for position- and time-resolved photoelectron and ion imaging was demonstrated. 

We demonstrate the application of such an in-vacuum pixel detector for bio-molecular MSI on an ion microscope for the first time. This detector concept comprises a fully-integrated, hybrid solid state pixel detector of the Medipix/ Timepix detector family. The presented assembly consists of a chevron MCP followed by 4 Timepix chips, which on the pixel level return (1) the impact position of a particle and (2) the TOF of this particle. The implementation of a Timepix detector for MSI on a TOF microscope mass spectrometer removes the need for mass-selection using an electrostatic blanker as used previously. Mass-resolved images are acquired during a single imaging experiment with this new detector setup.

Mass-resolved ion images from Timepix measurements of peptide and protein standards demonstrate the capability to return both mass-spectral and localization information of biologically relevant analytes from matrix-assisted laser desorption ionization (MALDI) on a commercial ion microscope.

Our work highlights the additional information that an MCP-Timepix detector assembly offers the MSI community. The comparison of Timepix acquired spectra to “standard” acquisition techniques reveals that the Timepix delivers an order of magnitude greater detectable range than an ADC and returns mass spectra for MCP gains from 4 · 10⁵ to 6.5 · 10⁶. Features of interest are the detector dynamic range, the available mass range, the detection homogeneity, the ability to detect single particles, the ability to resolve isotopes and the detector response to varying MCP gains.

On the TRIFT ion microscope, the Timepix assembly can detect oligomers of the protein ubiquitin up to 78 kDa.The mass spectrum of the protein ubiquitin as acquired with the Timepix detector on our ion microscope. The mass range up to 78 kDa is covered.