HMB-45 antibody demonstrates melanosome specificity by immunoelectron microscopy.

The intracellular localization of antigenic sites recognized by the monoclonal antibody HMB-45 was investigated in melanomas of the choroid and skin by postembedding immunoelectron microscopy. Antigenic sites were detected by a three-step procedure, consisting of incubating sections with the monoclonal HMB-45 antibody (protein G affinity-purified ascites from Enzo Diagnostics Inc or tissue culture supernatant from Dako Corp), followed by incubation with an affinity-purified rabbit anti-mouse IgG and finally with protein A-gold complex. Gold particles, indicative of HMB-45 immunoreactivity, were restricted to melanosomes in the malignant melanocytes. Early stages in melanosome formation (stages I through III) were most intensely stained, while late-stage melanosomes (stage IV) were only sparsely labeled or not stained at all. Melanophages adjacent to a cutaneous melanoma showed intense immunoreactivity in the cytoplasm and especially over electron-dense portions of lysosomes with the HMB-45 antibody from Enzo. In marked contrast, only very sparse labeling was detected over melanophages using a similar concentration of the HMB-45 antibody from Dako. Subsequently, when the Enzo antibody was diluted 40 times above the recommended working dilution, most of the melanophage staining disappeared, while melanocyte-specific staining was maintained. Immunolabeling of melanosomes with HMB-45 was drastically reduced or absent following section pretreatment with neuraminidase, confirming an earlier report that the HMB-45 antigen is partially composed of sialic acid. Our immunoelectron microscopic results show that HMB-45 antibody specifically stains melanosomes, rather than diffuse cytoplasmic antigen, as described by light microscopic immunohistochemical analysis, thus explaining its specificity for melanocytes. In addition, the elimination of HMB-45 immunoreactivity by neuraminidase pretreatment supports the idea that sialylation of antigen is crucial to HMB-45 binding, and suggests that the absence of staining in normal adult melanocytes, dermal nevi, and other melanocytic lesions may be a result of differential sialylation.

Changes in thyroid function tests and sex hormone binding globulin associated with treatment by gonadotropin.

Increasing levels of E2 in gonadotropin-treated women stimulated hepatic synthesis of SHBG and TBG, and consequently increases in T4 concentration. Nevertheless, unchanged FTI and TSH suggested that a euthyroid state was maintained. The temporal patterns for the rise in serum concentrations of TBG and SHBG during gonadotropin therapy suggest that the synthesis of these proteins by the liver has different sensitivities to E2.

Anaplastic thyroid tumors. Immunohistology.

Using an avidin-biotin immunoperoxidase technic, the authors studied 29 anaplastic thyroid tumors (ATTs) to determine the frequency of hormonal (thyroglobulin--TG; calcitonin--CT), epithelial (epithelial membrane antigen, monoclonal keratin), or sarcoma (desmin; alpha-1-antichymotrypsin--ACT; vimentin) markers. Their results indicate that 27% of ATTs stain for TG and none for CT. Fifty-five percent showed epithelial markers, 48% marked for ACT, and 47% for vimentin. Coexpression of keratin and vimentin was found in 39% of cases tested. The expression of the tested antigens did not correlate significantly with histologic pattern (epithelial vs. "sarcomatous"). Of note is the fact that 30% of the ATTs the authors tested expressed none of the markers they examined, indicating total lack of differentiation.