Evaluation of Long-Term Cryostorage of Brain Tissue Sections for Quantitative Histochemistry

Storage of tissue sections for long periods allows multiple samples, acquired over months or years, to be processed together, in the same reagents, for quantitative histochemical studies. Protocols for freezer storage of free-floating frozen sections using sucrose with different additives have been reported and assert that storage has no effect on histochemistry, but no quantitative support has been provided. The present study analyzed the efficacy of long-term storage of brain tissue sections at −80C in buffered 15% glycerol. To determine whether histochemical reactivity is affected, we analyzed 11 datasets from 80 monkey brains that had sections stored for up to 10 years. For processing, sections from multiple cases were removed from storage, thawed, and batch-processed at the same time for different histochemical measures, including IHC for neuronal nuclear antigen, parvalbumin, orexin-A, doublecortin, bromodeoxyuridine, the pro-form of brain-derived neurotrophic factor, and damaged myelin basic protein as well as a histochemical assay for hyaluronic acid. Results were quantified using stereology, optical densitometry, fluorescence intensity, or percent area stained. Multiple regression analyses controlling for age and sex demonstrated the general stability of these antigens for up to a decade when stored in 15% glycerol at −80C.

Giant Cell Granuloma of the Temporal Bone: A Case Report With Immunohistochemical, Enzyme Histochemical, and In Vitro Studies

A case of giant cell granuloma (GCG) that occurred in the right temporal bone is reported. The lesion showed histologic features identical to GCG. The multinuclear giant cells (MGCs) in the lesion showed strong reactivity with CD68, but patchy staining for myeloid/histiocyte antigen, α-1-antitrypsin, α-1-antichymotrypsine, and lysozyme. Activity of tartrate-resistant acid phosphatase was also consistently detected in the MGCs. Some of the mononuclear cells of the lesion exhibited similar immunocytochemical and histochemical reactivity as the MGCs. Ki-67 staining, however, was only detected in the mononuclear cells. The MGCs isolated from the lesion presented characteristic morphology of osteoclasts and possessed the ability to excavate bone in vitro. Thus, the MGCs in GCG appeared to express both macrophage- and osteoclast-associated phenotypes. The mononuclear cells were the major proliferative elements in the lesion and a subpopulation of these cells may represent precursors of the MGCs.

Determinant factors for the formation of the calcium oxalate minerals, weddellite and whewellite, on the surface of foliose lichens

The factors influencing the predominance of one of the two mineral forms of calcium oxalate (CO), the monohydrated whewellite (COM) and the di-hydrated weddellite (COD), forming the pruina of the upper cortex of lichens, have been investigated through a simple, sensitive histochemical assay: toluidine blue O (TBO), a metachromatic staining test. The differential reactivity of 43 thalli of 17 pruinose foliose species, supplemented by X-ray diffraction analysis and observations with polarizing and scanning electron microscopy, suggests that the histochemical reactivity of hyphal walls and cementing substances of the upper cortex are related to the density of anionic charges. These factors are probably due to the occurrence of polyuronic acid substances that strongly affects the mineralization of CO. Di-hydrated wedellite is always associated with TBO metachromatic reactivity, and COM with orthochromatic reactivity. When the material has an ambiguous ortho/metachromatic reactivity, COD and COM may occur together. This study presents the first experimental evidence that in lichens CO biomineralization is at least partially biologically controlled.

Histochemical Reactivity of Normal, Metaplastic, and Neoplastic Tissues to α-Linked N-Acetylglucosamine Residue-specific Monoclonal Antibody HIK1083

Monoclonal antibody (MAb) HIK1083, which is obtained by immunizing mice with a preparation of rat gastric mucins, has been shown to bind specifically to α-linked N-acetylglucosamine (α-GlcNAc). We investigated the specificity of MAb HIK1083 by immunostaining normal human organs, mucinous metaplasia of human pancreas, adenocarcinomas of human stomach, pancreas, and colon, and normal rat organs. The specificity was investigated by making comparisons with (a) a stain that labels Class III concanavalin A (ConA)-reactive mucin (Class III mucin), i.e., paradoxical ConA (PCS), and (b) staining with horseradish peroxidase (HRP)-conjugated Griffonia simplicifolia agglutinin II (GSA-II). In normal human and rat organs and in mucinous metaplasia of human pancreas, immunostaining with MAb HIK1083 and PCS showed similar specificities for mucins in glandular mucous cells. In adenocarcinoma of stomach and pancreas, GSA-II showed the most widespread positivity, PCS showed the least, and MAb HIK1083 showed a reactivity between those two extremes. Colon adenocarcinomas were labeled only with GSA-II. These results demonstrate that MAb HIK1083 could be a useful screening tool for Class III mucin in normal, metaplastic, and carcinoma tissues, and that the α-GlcNAc residue is one of the specific sugar residues found in Class III mucin.