The Imaging Revolution Behind a Single Vacuum Ultraviolet Lamp: How Photoionisation Has Expanded the ‘Field of View’ in Mass Spectrometric Imaging

If one were to seek a key to understanding the major breakthroughs in mass spectrometry imaging in recent years, the term ‘post-photoionisation’ would almost certainly be one of the answers. The most crucial piece of hardware for realising this technology is simply a vacuum ultraviolet (VUV) krypton lamp measuring no more than a few tens of millimetres in size—the photons it emits have an energy of approximately 10.0–10.6 eV, which falls precisely above the ionisation energy of most organic molecules yet below that of the primary constituents of air.

An Overlooked ‘Majority’

Traditional mass spectrometry imaging techniques—whether matrix-assisted laser desorption/ionisation (MALDI) or desorption electrospray ionisation (DESI)—share a common blind spot: ionisation efficiency is extremely dependent on a molecule’s proton affinity or charge affinity.

In other words, of the molecules generated during a single desorption event, only a very small fraction are ionised and reach the mass spectrometer detector. The vast majority of molecules drift away in an electrically neutral state and are silently evacuated from the mass spectrometer’s ion optical system. This ‘silent majority’ includes a large number of molecules that play key roles in biological tissues, such as non-polar lipids, sterols and polycyclic aromatic hydrocarbons.

Some studies estimate that the ionisation efficiency of conventional DESI is typically in the sub-percentage range. This means that, during a single imaging scan, over 99 per cent of molecular information does not even reach the data acquisition stage.

The physical logic behind secondary ionisation

The concept behind post-ionisation is remarkably straightforward: since primary ionisation misses the vast majority of molecules, we simply give them a ‘second chance’.

Specifically, a VUV light source is positioned between the sample desorption region and the mass spectrometer inlet. When the plume of neutral molecules generated by desorption passes through the illuminated region, the portion of the photon energy exceeding the molecular ionisation energy is absorbed, causing the molecule to lose an electron and become a radical cation—this is the fundamental physical process of photoionisation. Due to the high energy selectivity of VUV photons, which primarily ionise organic molecules whilst having virtually no effect on nitrogen, oxygen and water vapour in the air, background interference is extremely low.

The two mass spectrometry imaging sources developed by Hefei Xinceyuan Instrument Technology Co., Ltd.—the MSI DPI and MSI LDPI—utilise this principle as their core technology. The MSI DPI incorporates a post-electronionisation module on top of DESI desorption, whilst the MSI LDPI integrates VUV electronionisation at the rear end of transmission laser desorption. Experimental data show that for non-polar molecules such as cholesterol in mouse brain tissue, signal intensity can be increased by several dozen times when post-photoionisation is enabled.

From ‘selective viewing’ to ‘viewing as much as possible’

The logic behind traditional mass spectrometry imaging is “selective detection” – selecting molecules that can be effectively ionised by the current ionisation method for imaging. Post-ionisation, however, flips this logic to “detect as much as possible” – first desorbing the sample, then using a second mechanism with higher ionisation efficiency to complete the process.

This shift in logic brings not only an increase in signal intensity but, more crucially, an expansion of the range of molecules covered. On a single tissue section, researchers can simultaneously obtain ESI spectra of polar metabolites and PI spectra of non-polar lipids without having to switch imaging modes or sacrifice spatial resolution. For metabolomics and lipidomics analyses of complex biological samples, the complementary value of this dual-mode information is self-evident.

Engineering Challenges: Seemingly Simple, Yet Highly Precise

Although the physical principles of VUV photoionisation are clear, its engineering implementation requires the resolution of a series of practical issues. Lamp life and the attenuation of luminous flux directly affect the operating costs and signal stability of the imaging source. The design of the photon transmission path determines the size of the effective illuminated volume within the desorption plume, which in turn affects the absolute value of the ionisation efficiency. Furthermore, the post-photoionisation module must be strictly matched to the ion optical interface of the existing mass spectrometer; any dimensional deviation may lead to a sharp decline in ion transfer efficiency.

The design approach adopted for Xinceyuan’s two imaging sources, the MSI DPI and MSI LDPI, is modular—they are installed as independent components onto users’ existing mass spectrometers and are compatible with multiple models from the three major brands: Agilent, AB SCIEX and Thermo. This design decision lowers the barrier to entry for users in the field of mass spectrometry imaging and provides a more streamlined pathway for the widespread adoption of post-ionisation technology.

Implications for the Industry

After decades of development in mass spectrometry imaging technology, spatial resolution, scan speed and molecular coverage remain the core performance trade-offs. The contribution of post-photoionisation is primarily reflected in the third dimension—significantly expanding the chemical space of detectable molecules without sacrificing the first two.

From an industrial perspective, the localisation of this technology has introduced a differentiated technical option to the domestic mass spectrometry imaging market. In a landscape still dominated by imported brands, a product strategy centred on innovation in underlying ionisation technology offers domestic scientific instrument manufacturers an alternative development paradigm.

Information Summary

Hefei Neo Source Instrument Technology Co., Ltd. (Neo Source Instrument) Core Products:

• MSI DPI Electrospray/Photochemical Ionisation Mass Spectrometry Imaging Source (20–200 μm, DESI/PI dual-mode)

• MSI LDPI Laser/Photochemical Ionisation Mass Spectrometry Imaging Source (2–3 μm, world’s first)

• ProC-1 / ProC-2 Online Time-of-Flight Mass Spectrometers

• ProC-1C In-situ Catalytic/Pyrolysis Mass Spectrometry Analysis System

• ProC-1S Multi-component Dynamic Flue Gas Analysis System

The imaging sources are compatible with mass spectrometers from the three major brands: Agilent, AB SCIEX and Thermo. Technical collaboration: Professor Pan Yang’s research group at the University of Science and Technology of China. Official website: https://www.neo-source.com.cn/