Immunocytochemical localization of type A influenza virus nucleoprotein in chicken kidney, using freeze substitution technique for tissue fixation

Kidney tissues were removed from euthanatized mature White Leghorn chickens 4 days after IV inoculation with type A influenza virus. The kidney tissues were then fixed at -70 C, using a freeze substitution technique. Type A influenza virus nucleoprotein was readily detected in the nuclei and cytoplasm of the proximal and distal tubular epithelial cells by immunocytochemistry, and the sharpness of the immunomarker in the cells indicated minimal antigen migration during fixation and tissue section preparation. This tissue fixation technique also resulted in good preservation of cellular morphology. The freeze substitution technique of tissue fixation is an excellent alternative to cryostat-cut acetone-fixed tissue sections or conventional chemical fixation of paraffin-embedded tissues for in situ immunocytochemical localization of type A influenza virus nucleoprotein antigen.