Bringing SEM and MSI Closer Than Ever Before: Visualizing Aspergillus and Pseudomonas Infection in the Rat Lungs

Area of Science:

• Pathology
• Microscopy
• Mass Spectrometry Imaging

Background:

• Frozen tissue sections are valuable for both structural and molecular analysis.
• Simultaneous structural and molecular imaging of infected lung tissue presents technical challenges.

Purpose of the Study:

• To develop a protocol for processing frozen rat lung tissue sections for scanning electron microscopy (SEM).
• To enable subsequent matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) on the same tissue sections.
• To demonstrate bimodal image fusion of SEM and MALDI-MSI data.

Main Methods:

• Developed a slow thawing and fixation procedure for frozen lung sections.
• Utilized graded ethanol dehydration and critical point drying for SEM sample preparation.
• Processed consecutive nonfixed sections for MALDI-MSI.
• Employed in-house software (CycloBranch) for bimodal image fusion.

Main Results:

• SEM imaging revealed well-preserved tissue structures, including bacterial cells and fungal hyphae.
• MALDI-MSI provided molecular biomarker maps of *Pseudomonas aeruginosa* infection.
• Successful bimodal image fusion of SEM and MALDI-MSI data was achieved.

Conclusions:

• The developed protocol effectively processes frozen rat lung tissue for both SEM and MALDI-MSI.
• This method allows for correlative structural and molecular analysis of infected lung tissue.
• Bimodal imaging provides comprehensive insights into host-pathogen interactions.