Optimized methyl green-pyronin Y staining for layer visualization in frozen mouse cerebellum

Abstract

Background

Conventional histological stains, such as hematoxylin and eosin (H&E) or toluidine blue O (TBO), have a limited ability to clearly delineate the layered architecture of the cerebellar cortex.

New method

We applied methyl green–pyronin Y (MGP) staining, which is traditionally used for nucleic acid differentiation, to frozen mouse cerebellar sections to enhance visualization of cortical layers and neuronal subtypes.

Results

MGP staining yielded strong contrast between cell types: Purkinje cells stained distinctly pink, while granule cells appeared green. This enabled clear identification of cerebellar lamination and neuronal distribution.

Comparison with existing methods

In H&E or TBO staining, Purkinje and granule cells are colored similarly, which obscures layer boundaries. Although immunohistochemistry is commonly used to distinguish these cell types, MGP staining provides a rapid, color-based distinction without the need for antibodies or fluorescence.

Conclusions

MGP staining provides a fast and cost-effective alternative for analyzing cerebellar tissue, enabling clear visualization of cortical layering and facilitating the morphological screening of cerebellar abnormalities.