Agarose infiltration improves morphology of cryostat sections of lung

BACKGROUND: Optimal morphology and immunohistochemistry of the lung requires the organ to remain fully distended. When the lung is prepared for frozen sections, especially during the stage of cryoprotection, liquid fixatives leak out and the lung tends to collapse. We sought to develop a new technique to improve the morphology of frozen sections of the lung while facilitating tissue handling and immunoreactivity. EXPERIMENTAL DESIGN: Human, fetal sheep and adult rat lungs were distended with 1% low temperature melting agarose and vessels were perfused with a 1% paraformaldehyde solution. RESULTS: The agarose infiltration permitted easy handling of the lung tissue, and maintained lung architecture during cryoprotection. Agarose infiltration and paraformaldehyde fixation provided excellent morphology, that was comparable to paraffin-embedded tissue. The structural preservation was especially noticeable with human lung that remained well distended with the agarose technique. Because of the mild fixation and good morphology, precise localization of antigenic sites was possible for the surfactant proteins SP-A, SP-C and SP-D, cytokeratin, and alveolar macrophage markers. CONCLUSIONS: This procedure can improve special fixation and embedding protocols for lung immunocytochemistry and immunofluorescence.