MALDI-IMS experiments were performed as previously described (14 (link)). In brief, BxPC3 tumor xenografts were
removed and flash-frozen in liquid nitrogen before storage at
−80°C. Frozen tissue samples were then cut into 12 μm thick
sections using a cryostat (Leica Microsystems, Buffalo Grove, IL). These frozen
sections were then thaw-mounted onto gold-coasted, stainless steel MALDI target
plates. Matrix solution (2’,4’,6’-trihydroxyacetophenone,
20 mg/mL in 60:40 methanol:water with 0.1% trifluoroacetic acid) was manually
applied to tissues using a glass nebulizer. MALDI mass spectra were acquired
using a linear ion trap (LTQ XL) mass spectrometer (Thermo Fisher Scientific) in
MS/MS mode. MS/MS was performed on the protonated parent ion of gemcitabine (m/z
264) and full product ion spectra were obtained, allowing GEM to be
differentiated by pseudo-selected reaction monitoring. The main fragment ion of
gemcitabine at m/z 112 was utilized for image reconstruction. Spectra were
obtained for each section at 150 μm spatial resolution. Images were
generated using ImageQuest software (Thermo Fisher Scientific) by analyzing the
main fragment ion intensity as a function of the position over the tissue
surface. For statistical comparisons, average spectra were generated over each
tissue section and the intensities of m/z 112 were exported for further
comparisons.
removed and flash-frozen in liquid nitrogen before storage at
−80°C. Frozen tissue samples were then cut into 12 μm thick
sections using a cryostat (Leica Microsystems, Buffalo Grove, IL). These frozen
sections were then thaw-mounted onto gold-coasted, stainless steel MALDI target
plates. Matrix solution (2’,4’,6’-trihydroxyacetophenone,
20 mg/mL in 60:40 methanol:water with 0.1% trifluoroacetic acid) was manually
applied to tissues using a glass nebulizer. MALDI mass spectra were acquired
using a linear ion trap (LTQ XL) mass spectrometer (Thermo Fisher Scientific) in
MS/MS mode. MS/MS was performed on the protonated parent ion of gemcitabine (m/z
264) and full product ion spectra were obtained, allowing GEM to be
differentiated by pseudo-selected reaction monitoring. The main fragment ion of
gemcitabine at m/z 112 was utilized for image reconstruction. Spectra were
obtained for each section at 150 μm spatial resolution. Images were
generated using ImageQuest software (Thermo Fisher Scientific) by analyzing the
main fragment ion intensity as a function of the position over the tissue
surface. For statistical comparisons, average spectra were generated over each
tissue section and the intensities of m/z 112 were exported for further
comparisons.
