Next Generation Histology-Directed Imaging Mass Spectrometry Driven by Autofluorescence Microscopy

Histology-directed
imaging mass spectrometry (IMS) is a spatially
targeted IMS acquisition method informed by expert annotation that
provides rapid molecular characterization of select tissue structures.
The expert annotations are usually determined on digital whole slide
images of histological stains where the staining preparation is incompatible
with optimal IMS preparation, necessitating serial sections: one for
annotation, one for IMS. Registration is then used to align staining
annotations onto the IMS tissue section. Herein, we report
a next-generation histology-directed platform implementing IMS-compatible
autofluorescence (AF) microscopy taken prior to any staining or IMS.
The platform enables two histology-directed workflows, one that improves
the registration process between two separate tissue sections using
automated, computational monomodal AF-to-AF microscopy image registration,
and a registration-free approach that utilizes AF directly to identify
ROIs and acquire IMS on the same section. The registration approach
is fully automated and delivers state of the art accuracy in histology-directed
workflows for transfer of annotations (∼3–10 μm
based on 4 organs from 2 species) while the direct AF approach is
registration-free, allowing targeting of the finest structures visible
by AF microscopy. We demonstrate the platform in biologically relevant
case studies of liver stage malaria and human kidney disease with
spatially targeted acquisition of sparsely distributed (composing
less than one tenth of 1% of the tissue section area) malaria infected
mouse hepatocytes and glomeruli in the human kidney case study.