RESUMO
Cryostat sectioning is a popular but labor-intensive method for preparing histological brain sections. We have developed a modification of the commercially available CryoJane tape collection method that significantly improves the ease of collection and the final quality of the tissue sections. The key modification involves an array of UVLEDs to achieve uniform polymerization of the glass slide and robust adhesion between the section and slide. This report presents system components and detailed procedural steps, and provides examples of end results; that is, 20 µm mouse brain sections that have been successfully processed for routine Nissl, myelin staining, DAB histochemistry, and fluorescence. The method is also suitable for larger brains, such as rat and monkey.
Assuntos
Encéfalo/ultraestrutura , Crioultramicrotomia/métodos , Microtomia/métodos , Coloração e Rotulagem/métodos , 3,3'-Diaminobenzidina/química , Animais , Crioultramicrotomia/instrumentação , Haplorrinos , Ensaios de Triagem em Larga Escala , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Microscopia de Fluorescência , Microtomia/instrumentação , Bainha de Mielina/ultraestrutura , Corpos de Nissl/ultraestrutura , Ratos , Ratos Long-Evans , Coloração e Rotulagem/instrumentaçãoRESUMO
A major challenge in the histological sectioning of brain tissue is achieving accurate alignment in the standard coronal, horizontal, or sagittal planes. Correct alignment is desirable for ease of subsequent analysis and is a prerequisite for computational registration and algorithm-based quantification of experimental data. We have developed a simple and low-cost technique for whole-brain cryosectioning of rodent brains that reliably results in a precise alignment of stereotactic coordinates. The system utilises a 3-D printed model of a mouse brain to create a tailored cavity that is used to align and support the brain during freezing. The alignment of the frozen block is achieved in relation to the fixed edge of the mold. The system also allows for two brains to be frozen and sectioned simultaneously. System components, procedural steps, and examples of the end results are presented.