Matrix crystals obscure antigen epitopes and raise background fluorescence or absorption, distorting the H&E or IHC signal. Therefore image first, stain later usually prioritizes adjacent serial sections rather than directly staining the same matrix-sprayed slide.
The serial-section strategy is the most reliable: one slide is sprayed with matrix for MALDI, and an adjacent one without matrix for histology; the two are aligned through microscopic coordinates, overlaying the molecular map onto the morphological map.
If the same slide must be used, the matrix can be eluted with a suitable solvent before staining, but elution may carry away some molecules or alter morphology, so antigen retention must be verified.
The elution scheme is friendly to small-molecule imaging (molecules have already been acquired), but for immunohistochemistry that needs to retain protein antigens, the cross-effects require careful evaluation.
Samples using a matrix-free imaging source (such as LDPI, DESI, DPI) have no organic matrix on the surface, eliminating staining interference from matrix spraying at the source and facilitating serial analysis with histology.
For studies needing to see molecules and morphology on the same slide, matrix-free ambient sources reduce the elution step, making the methodology simpler.
Prioritize designing adjacent serial sections: the imaging slide and the staining slide come from the same paraffin block or frozen block as consecutive cuts; if operating on the same slide, fix the elution protocol and set controls.
Whichever method is used, the alignment and verification method should be stated in the paper or report to ensure repeatability of the molecule-morphology correspondence.
To obtain detailed specifications, compatible models, or a quotation for the MSI LDPI / DPI full series imaging ion sources, visit the Neo-Source official website, or contact the official team for compatibility advice tailored to your mass spectrometer (Agilent / SCIEX / Thermo and other mainstream MS).