Automatic Sprayer for MALDI Imaging iMLayer AERO

Importance of MALDI Imaging

Matrix-assisted laser desorption/ionization (MALDI) imaging has become a cornerstone technique in spatially resolved molecular analysis of biological tissues. High sensitivity and spatial resolution are essential for mapping biomolecules such as lipids, peptides and small metabolites in situ. The iMLayer™ AERO system addresses common challenges in MALDI sample preparation by automating pretreatments and delivering uniform matrix coatings, improving reproducibility and throughput in research and QC laboratories.

Objectives and Study Overview

This application note introduces the iMLayer AERO automatic sprayer designed for MALDI imaging workflows. Key goals include demonstrating how controlled matrix deposition enhances signal intensity and spatial fidelity, automates complex pretreatment steps (e.g. derivatization, enzyme digestion) and integrates easily with downstream mass spectrometry imaging (MSI) analysis.

Methodology and Workflow

The iMLayer AERO workflow comprises four main stages:

• Derivatization or enzyme treatment: optional chemical modification to improve ionization efficiency (e.g., Girard’s reagent T for steroids, trypsin digestion of proteins).
• Matrix spraying: a programmable nozzle deposits fine crystals of DHB, 9-AA or CHCA in multiple laminating passes for uniform coverage.
• MSI acquisition: prepared slides are analyzed by a MALDI-MS instrument, producing spatially resolved mass spectra.
• Data analysis: specialized software reconstructs ion images and quantifies analyte distributions.

The system replaces manual pipetting and hand spraying, reducing user dependence and variability.

Used Instrumentation

A summary of critical hardware and software components:

• iMLayer AERO sprayer with adjustable nozzle-to-sample distance (5–10 cm) and motorized sample stage (speed up to 70 mm/s).
• Humidity-controlled spray chamber with N₂ purging to ensure consistent atmospheric conditions.
• Automated rinse mechanism to prevent nozzle clogging and maintain stable reagent flow.
• Control software with wizard-guided setup and preset methods for error-free operation.
• Optional two-step vapor deposition unit (iMLayer) for ultra-fine matrix films (1 µm) enabling spatial resolution down to 5–10 µm.

Main Results and Discussion

Comparative tests show that the automated spray protocol matches or exceeds the sensitivity of manual spraying while delivering significantly improved homogeneity and reproducibility. The two-step vapor deposition method further refines spatial resolution, producing images with minimal blur compared to hand-sprayed samples. Derivatization experiments on testosterone spots confirmed enhanced ion yields and more reliable quantification.

Benefits and Practical Applications

The iMLayer AERO platform offers:

• High reproducibility: automated environmental control and nozzle cleaning minimize variability.
• Enhanced sensitivity: fine crystal layers boost ion signal intensity.
• Improved spatial resolution: two-step coating delivers sharper images for tissue sections.
• Versatility: supports multiple matrices, derivatization and enzyme digestion protocols.
• Time savings: streamlined workflows reduce hands-on preparation and training requirements.

Applications span pharmaceutical research, biomarker discovery, clinical pathology and quality control in industrial settings.

Future Trends and Potential Applications

Advancements in automated sample preparation will continue to drive improvements in MSI throughput and data quality. Integration with high-speed imaging mass spectrometers and AI-based image analysis promises real-time tissue profiling. Further development of novel derivatization chemistries and matrix materials may expand the range of detectable analytes, supporting metabolomics and single-cell studies.

Conclusion

The iMLayer AERO system represents a significant step forward in MALDI imaging sample preparation. By automating complex pretreatments and delivering reproducible matrix coatings, it enhances both the sensitivity and spatial fidelity of MSI experiments. This platform empowers researchers and analytical laboratories to obtain high-quality molecular images with reduced hands-on time and improved consistency.