A new method of scanning electron microscopy for imaging biological tissues

Area of Science:

• Materials Science
• Microscopy
• Histology

Background:

• Scanning electron microscopy (SEM) is limited for histological samples due to surface-sensitive secondary electron imaging.
• Conventional SEM imaging distorts the actual structure of processed and cut histological sections.
• Existing methods using backscattered electrons (BSE) require non-transparent substrates, preventing correlative light microscopy.

Purpose of the Study:

• To develop a method for imaging histological sections in SEM with resolution superior to light microscopy.
• To enable correlative imaging of the same histological section using both light microscopy and SEM.
• To overcome the limitations of surface-based imaging in SEM for biological tissues.

Main Methods:

• Utilizing the cathodoluminescence (CL) signal emitted by standard glass microscope slides.
• Employing electron flux through the histological section as a measure of image signal intensity.
• Imaging 7-micrometer thick histological sections mounted on glass slides using SEM at 5-20 keV.

Main Results:

• Achieved SEM imaging resolution exceeding that of conventional light microscopy for histological sections.
• Demonstrated that glass slide cathodoluminescence is proportional to the electron flux transmitted through the specimen.
• Enabled correlative imaging of histological sections mounted on transparent glass slides.

Conclusions:

• Cathodoluminescence imaging of glass-mounted histological sections in SEM offers a valuable alternative to traditional methods.
• This technique provides higher resolution than light microscopy and allows for direct correlation with light microscopic images.
• The method overcomes the limitations of surface-dependent imaging in SEM for histological samples.