Feasibility of Biological Mass Spectrometry Imaging and Depth Profiling by Ultrashort Laser Pulses

Biological mass spectrometry (MS) imaging serves to visualize the spatial distribution of molecules directly from mammalian, or plant tissue, and microbial biofilms. Maps of hundreds of molecules from such samples are welcome in many areas of biomedical research. MS imaging faces three challenges: molecular tomography to allow 3D molecular mapping; overcoming of the current limits of spatial resolution; minimization of sample preparation. The work in the present thesis evaluated the fundamental feasibility of using the near-IR ultrashort laser pulses both as a microprobe for high resolution 2D MS imaging and for in situ depth profiling. It was shown that sub-100 fs, ~800 nm laser pulses can be used to ablate ~10 μm thick topmost layer from the surface of ~100 μm thick bacterial biofilms leaving behind underlying layer exposed for subsequent MS analysis. Similarly, it was found that fs laser ablation can remove several tens of micrometers from the surface of an eye lens tissue with precision of ±5 μm. MS analysis from intact and ablated biofilms indicated no chemical degradation of an antibiotic doped therein, while MS analysis of cholesterol, phospholipids, peptides, and various unidentified endogenous species from the intact and ablated eye lens tissue revealed only minor chemical damage caused by fs laser ablation. Finally, ultrashort pulse laser desorption coupled with postionization by 10.5 eV laser pulses was used to analyze simulated and actual biological substrates spiked with thermometer molecule 4-chlorbenzylpyridinium (CBP) chloride. Internal energy (Eint) of CBP ion produced under MS imaging-like experimental conditions corresponding to ~30 µm spatial resolution was found to be comparable to that of established “soft” MS imaging techniques such as MALDI-MS.