Immunohistochemical and ultrastructural localization of tissue polypeptide antigen in human ovarian tumours

Summary Forty specimens of benign and malignant ovarian tumours were studied for localization of tissue polypeptide antigen (TPA) at light and electron microscopic levels by an indirect immunoperoxidase technique. Of the 30 ovarian carcinomas, 23 (77%) were positive and 7 (23%) were negative for TPA, while of the 10 benign ovarian tumours 3 (30%) were positive and 7 (70%) were negative. Positive reaction did not correlate with the tumour grade. Of the 10 patients with metastasis, 8 (80%) had positive tumours. Staining for TPA was observed at the intraluminal cell surfaces and peripheral cell membranes. The ultrastructural localization of TPA revealed electron-dense reaction products at the cell surface and microvillous surfaces. These results provide confirmatory and supplementary evidence to support the previous findings of TPA in the serum and suggest that testing for TPA in ovarian tumors has a limited prognostic importance and a poor diagnostic value. The surface property of TPA suggests that the cell membrane is involved in secretion and probably synthesis of TPA.     

Immunohistochemical localization of CA 125 antigen in formalin-fixed paraffin sections of ovarian tumors with the use of Pronase

The OC 125 monoclonal antibody was used to localize CA 125 antigen in routine paraffin sections of ovarian tumors with the use of a modified avidin-biotin-peroxidase complex (ABC) technic. Pretreatment of the paraffin sections with Pronase allowed subsequent detection of CA 125 antigen. OC 125 stained 4 (80%) of 5 benign and borderline serous ovarian tumors, 12 (86%) of 14 serous adenocarcinomas, and 3 (23%) of 13 benign and malignant mucinous ovarian tumors. The pattern of distribution of CA 125 antigen was mostly at the intraluminal and peripheral cell surfaces, while intracytoplasmic staining also was seen. Overall, CA 125 antigen detectability rate in paraffin sections was found to be compatible with those reported in frozen sections. The method allows retrospective immunohistochemical examination of a large number of cases with ovarian tumors.     

Immunohistochemical localization of epidermal and Mallory body cytokeratin in undifferentiated epithelial tumors. Comparison with ultrastructural features

Twenty-one anaplastic tumors were studied by light microscopy (LM), immunoperoxidase staining using anti-epidermal cytokeratin (ECK) and anti-Mallory body cytokeratin (MBCK) antibodies, and electron microscopy (EM), to determine whether an epithelial origin could be confirmed. The tumors were derived from lung, stomach, colon, breast, uterus, kidney, bladder, and mesothelium. By LM, the tumors consisted of either large and polygonal, spindle or small, round cells. With immunoperoxidase staining, 11 (52%) of the anaplastic tumors were positive for ECK, positivity being either absent or only weak in the main tumor mass, but marked in areas of infiltration and metastases. In contrast, all of the anaplastic tumors were positive for MBCK in the main tumor mass, infiltrating areas, and metastases. In the case of adenocarcinomas, staining was either web-like or diffuse throughout the cytoplasm with concentration occurring at the cell surface, whereas in mesotheliomas, the staining was either diffuse or showed focal perinuclear accentuation. Twelve of 13 anaplastic tumors examined by EM showed epithelial features (desmosomes, tonofilaments, lumina, and/or microvilli). As controls, 21 non-epithelial tumors (five melanomas, eight sarcomas, and eight lymphomas) showed no reactivity with either cytokeratin antibody. These studies show that the epithelial nature of undifferentiated and poorly differentiated tumors can be confirmed by immunohistochemistry using anti-cytokeratin antibodies.     

Immunohistochemical localization of renin in renal tumors.

Immunoperoxidase staining for renin was performed with renal tumors, including juxtaglomerular (JG) tumor, Wilms' tumors, renal adenocarcinomas, renal oncocytomas, and cortical adenomas. Compared with the JG apparatus adjacent to the glomerulus, JG tumor cells were less darkly but diffusely stained for renin. One of five Wilms' tumors revealed more numerous renin-containing tumor cells than the adjacent renal cortex, whereas three of ten renal adenocarcinomas and two of three renal oncocytomas revealed only focally renin-positive tumor cell cytoplasms. None of six cortical adenomas were positive for renin. With available fresh tumor tissue, renin activity was studied by measuring newly formed angiotensin I by radioimmunoassay. JG tumor contained markedly elevated renin activity, whereas one Wilms' tumor and two renal adenocarcinomas contained no more than 2% of renin activity of the renal cortex, more than 50% of which was inactive renin. These findings suggest that the JG tumor elaborates enormous amounts of active renin, whereas other renal tumors produce lesser amounts of renin, more than half of which is inactive renin.     

Immunohistochemical localization of metallothionein in human thyroid tumors.

High levels of metallothionein (MT) are present in the developing mammalian liver; however, a remarkable decrease is observed during postnatal life after weaning. This developmental profile is similar to that of certain oncofetal gene products such as alpha-fetoprotein, which is used as a tumor marker. This study deals with the reexpression of MT genes in thyroid tumors. With an immunohistochemical method, the presence of MT was investigated in tissue sections of normal and neoplastic human thyroid glands. Tissue sections of 34 thyroid tumors and 10 normal human thyroid glands were studied by means of the peroxidase-antiperoxidase method. MT was localized in 31 of the thyroid gland tumors. MT was also present in two of the normal thyroid glands. These findings indicate that although high levels of MT are mainly found in the fetal liver, it may also be expressed actively in certain human thyroid neoplastic tissues, and occasionally in normal thyroid tissue.     

Immunohistochemical localization of neurofilament antigen in rat cerebellum

Summary The distribution of neurofilaments in the rat cerebellar cortex was studied by immunoperoxidase histochemistry using an antiserum raised against neurofilaments isolated from brain (anti-NF). In light microscope preparations, this antiserum selectively stained known neurofilament-containing structures. Staining was most intense in myelinated axons of the white matter and in the terminal branches of basket cell axons. No staining was apparent in either neuronal or glial cell bodies or in glial cell processes. These findings were confirmed in electron microscopic preparations of the same material. Neurofilaments stained by the antiserum were abundant in basket cell axons and also occurred in small bundles in mossy fibre terminals. Adjacent microtubules were not stained by the antiserum. There was no evidence of stained cytoplasmic filaments in glial cell processes. Thus it appears that neurofilaments contain unique antigens which do not occur in either microtubules or in glial cytoplasmic filaments. The antiserum did not induce staining of synaptic junctional structures, a result which contradicts previous suggestions that neurofilaments are structural components of synaptic densities.     

Immunohistochemical localization of survivin in serous tumors of the ovary.

The aim of this study was to determine the immunohistochemical distribution of survivin in benign, borderline, and malignant serous tumors of the ovary. Survivin was localized by an indirect immunoperoxidase method in 42 cases of serous tumors of the ovary (15 cystadenomas, 15 borderline tumors, and 12 cystadenocarcinomas). Nuclear staining and cytoplasmic staining were separately scored. Cytoplasmic staining was detected in 27% of adenomas/borderline tumors and in 58% of carcinomas. Nuclear staining was detected in 87% of adenomas/borderline tumors but in only 42% of carcinomas. Although the differences in the intensity of cytoplasmic staining between adenomas and borderline tumors versus carcinomas were not significant, the differences in the intensity of nuclear staining between low-grade versus malignant tumors were significant. These findings suggest that survivin is widely expressed in benign, borderline, and malignant serous tumors but that nuclear localization of survivin is more common in benign or borderline tumors than in malignant serous tumors of the ovary. The molecular mechanisms that determine the subcellular distribution of this protein may reflect the role of survivin in the regulation of apoptosis during the processes of malignant transformation.     

The ultrastructural localization of keratin proteins and carcinoembryonic antigen in malignant mesotheliomas.

The immunoultrastructural localization of keratin proteins and carcinoembryonic antigen (CEA) in mesothelioma cells was accomplished with a low-temperature embedding colloidal gold technique. The keratin antiserum labeled only intermediate filaments. These filaments surrounded the cytoplasmic organelles and were inserted into desmosomes. The CEA antiserum labeled cytoplasmic vesicles and droplets. No definite labeling of microvilli was observed.     

Immunohistochemical characterization of ovarian borderline tumors of intestinal and mullerian types.

A panel of monoclonal antibodies (MAbs) including MOv2, MOv8, anti-CA 19-9, and anti-carcinoembryonic antigen (CEA), and a polyclonal antibody against CEA, was applied to three types of ovarian borderline tumors. The tumors included 19 intestinal-type mucinous borderline tumors (IMBTs), 22 endocervical-like mucinous borderline tumors (EMBTs), and 23 mixed-epithelial borderline tumors (MEBTs); the latter two tumors are of mullerian type. Statistically significant differences in the percentage of positive IMBTs compared to the mullerian tumors were seen for anti-CEA and MOv8; strikingly different staining patterns were also seen. IMBTs were more often and more diffusely CEA-positive than were the mullerian tumors; within the mullerian tumors, only one type of cell, the indifferent eosinophilic cell, was consistently CEA positive. The MAbs MOv2, MOv8, and anti-CA 19-9 showed more extensive positivity in the mullerian tumors than in the IMBTs. This study highlights the differences in cell types between IMBTs and these two types of mullerian borderline tumors.     

Immunohistochemical localization of S100 protein in skin tumors

We applied a peroxidase-antiperoxidase technique for S100 protein to 73 tumors of skin and skin adnexa. These included 15 eccrine tumors, 11 apocrine tumors, 18 tumors with differentiation toward hair, two sebaceous adenomas, one mixed tumor of the scalp, ten dermatofibromas, ten basal cell carcinomas, five squamous cell carcinomas, and one clear cell acanthoma. Consistent results were obtained. Occasional cells in eccrine tumors showed strong positive staining, as did the Langerhans' cells in the squamous cell carcinomas and the clear cell acanthoma. The cells of the apocrine tumors showed moderate to weak staining, and the tumors with differentiation toward hair, the sebaceous adenomas, and the mixed tumor of the scalp showed uniform negative staining, as did basal cell carcinomas and dermatofibromas.     

Immunohistochemical localization of WT1 in epithelial salivary tumors

The subcellular localization of WT1 is controversial and has received little attention in the epithelial tumors of salivary glands. Paraffin-embedded, surgical specimens from 80 salivary tumors were investigated by immunohistochemistry using a monoclonal, anti-WT1 antibody (6F-H2, Dako). Immunostaining was seen in 14/14 pleomorphic adenomas (PAs), 6/6 myoepitheliomas, 4/4 basal cell adenomas, 4/4 canalicular adenomas, 0/7 Warthin tumors, 0/1 oncocytoma, 1/6 acinic cell carcinomas (Cas), 0/11 mucoepidemoid Cas, 1/11 adenoid cystic Cas, 11/12 polymorphous low-grade adenocarcinomas (PLGAs), 1/2 Ca ex PA, 0/1 salivary duct Ca and 0/1 clear cell adenocarcinoma. Stained-cell subpopulations up to 90% were not uncommon in the benign tumors. Up to 80% of cells in PLGA could be stained. Staining was weak to intense and confined to the cytoplasm of preferentially non-luminal or adjacent to stroma cells. One adenoid cystic Ca showed nuclear staining. The results suggest that WT1 is often highly expressed in benign non-oncocytic salivary tumors whereas the malignant tumors show decreased expression, the exception being PLGA. The expression is usually cytoplasmic and associated with non-luminal cells. PLGA immunoreactivities could be useful in histological differential diagnosis.     

Immunohistochemical localization of human immunodeficiency virus p24 antigen in placental tissue.

As human immunodeficiency virus (HIV) infection spreads into the heterosexual population, perinatally acquired HIV infection will increase in incidence, and knowledge of the mechanism of this transfer is important. We have used immunoperoxidase techniques to detect HIV p24 antigen in formalin-fixed, paraffin-embedded placental tissue from nine known HIV serologically positive mothers. In four of these cases we have detected evidence or viral antigen in placental Hofbauer cells, vascular endothelium, or intermediate trophoblast. The implications for understanding the mode of transfer of infection to the fetus are discussed.     

Immunohistochemical localization of metal binding proteins in thyroid tissues and tumors

Positive immunohistochemical staining for three metal binding proteins, ceruloplasmin, lactoferrin, and transferrin, has been suggested to be a reliable diagnostic marker of malignant but not benign thyroid neoplasms. We tested this hypothesis on a series of 9 nodular hyperplasias, 17 follicular adenomas, 54 papillary carcinomas, 20 follicular carcinomas, and 3 anaplastic carcinomas of thyroid using formalin-fixed paraffin-embedded tissues. We found focal staining for ceruloplasmin and lactoferrin in approximately 25% of follicular adenomas examined; focal ceruloplasmin positivity was also seen in nonneoplastic tissues surrounding thyroid neoplasms. No staining for these markers was found in malignant neoplasms or hyperplasias. Transferrin was found in 55% of papillary carcinomas, but more follicular adenomas (20%) than follicular carcinomas (11%) were positive using this antiserum. These findings show that immunostaining for these antigens is not a reliable method to distinguish benign from malignant thyroid lesions of follicular cell origin.     

Immunohistochemical localization of the rabbit haemorrhagic disease viral antigen

Summary Immunohistochemical investigations were carried out to determine organ and cellular localization of the rabbit haemorrhagic disease viral antigen (RHDVA). It was found in certain parenchymal liver cells near the interlobular septs and in some macrophages and pseudoeosinophils of all studied organs and blood. Whereas in morphologically preserved hepatocytes and macrophages the RHDVA accumulated in the nuclei, in cells with disintegrated nuclei it was distributed throughout the cytoplasm.     

Reactivity of a monoclonal antibody with tissues and tumors from the human breast. Immunohistochemical localization of a new antigen and clinicopathologic correlations.

The reactions of a monoclonal antibody to the MCF7 breast cancer cell line were immunohistochemically studied on a variety of breast tumors, primary and metastatic, on mammary epithelium and on nonneoplastic breast lesions. A high proportion of positive reactions was observed in ductal, lobular, and tubular carcinomas as well as in mammary Paget's disease. Mucinous, medullary, and papillary carcinomas showed a low incidence of reactivity. Carcinomas with metaplasia, carcinoids, and nonepithelial breast tumors were unreactive with the antibody. Positive immunostaining was documented also in nodal and extranodal metastatic lesions. The staining of nodal metastases was correlated with the positive reaction of the primary tumor. Reactivity was widely distributed in normal breast epithelial cells and in benign breast lesions. Staining of nonneoplastic mammary epithelial was associated with reactivity of adjacent neoplastic tissues. Staining differences between nonneoplastic and neoplastic mammary tissues were related to the intensity and cytologic distribution of the labeling. Heterogeneous reactivity of morphologically similar cells was documented in nonneoplastic and neoplastic breast epithelial cells as well as in nodal and extranodal breast carcinoma metastases. Immunohistologically detectable antigen was not correlated with prognostic factors such as histologic grade or nodal status. A retrospective study of T1NO cases failed to substantiate any prognostic value for the reactivity of primary breast tumors with this monoclonal antibody.     

Immunohistochemical localization of factor X-like antigen in pancreatic islets and their tumours

Summary The authors have investigated by immunohistochemistry the distribution of factor X-like antigen in normal pancreatic islets and in a series of 46 pancreatic endocrine tumours. It was found that both glucagon-producing (A) cells and pancreatic polypeptide-producing (PP) cells are immunoreactive for the antigen. Benign glucagonomas and PP-omas presented the highest concentrations of immunoreactive material whose intracellular distribution was consistent with localization within cell secretory granules. Some benign insulinomas also presented factor X immunostaining in spite of the absence of the antigen in normal insulin-producing B cells. Although malignant tumours usually exhibited very low or no immunostaining, two of three malignant glucagonomas showed scattered, intensely immunoreactive cells. The factor X-like antigen identified in this study was found to differ from chromogranin A and B. The possible implications of the present findings for coagulative disorders associated with glucagonomas or diabetes are discussed.     

Immunohistochemical localization of murine stage-specific embryonic antigens in human testicular germ cell tumors.

Monoclonal antibodies raised against and/or recognizing stage-specific antigens on preimplantation mouse embryos and stem cells of murine teratocarcinoma were used to localize these antigens immunohistochemically on human testicular germ cell tumors. SSEA-1, the antigen found on mouse embryonal carcinoma (EC) cells and embryonic cells from the 8-cell stage embryo onward, including the fetal primordial germ cells, was detected on yolk sac carcinoma components of human tumors, but not on EC cells. SSEA-3, the antigen found on follicular ova, fertilized eggs, early cleavage stage embryonic cells, and visceral endodermal cells of the mouse embryo, but not on mouse EC cells, was detected on human EC cells. Both antigens were found on the cell surface of fetal testicular germ cells but not in the seminiferous tubules of adult human testes. These data point out differences between human and murine EC cells suggesting that human EC cells correspond developmentally to a less mature embryonic cell than the murine EC cells. The possible histogenesis of human germ cell tumors from primordial and/or fetal germ cells is briefly discussed.     

Immunohistochemical localization of Trk receptor protein in pediatric small round blue cell tumors.

Expression of Trk protein has been documented by Northern analysis in neuroblastomas with good prognosis. To localize the expression of this protein at the cellular level within individual tumors, we adapted a recently characterized pan-Trk antibody for use in formalin fixed, paraffin-embedded tissue. We have examined a group of small round blue cell tumors occurring in children, including both high and low stage neuroblastomas, to assess the presence or absence of Trk expression and its cellular localization. Positive staining for Trk protein was observed in four of four low stage (good prognosis) neuroblastomas, five of five primitive neuroectodermal tumors/Ewing's sarcoma, five of five rhabdomyosarcomas, and no lymphomas. Within the neuroblastomas, expression of Trk protein was most striking in ganglion cells, in which positive cytoplasmic staining was demonstrated regardless of tumor stage. The latter observation may lend further insight into the pathobiology of this malignant childhood tumor.     

Immunohistochemical localization of placental-like alkaline phosphatase in testis and germ-cell tumors using monoclonal antibodies.

Six monoclonal antibodies raised against the human placental alkaline phosphatase (ALP) recognizing distinct antigenic determinants on the surface of this isozyme were used for immunohistochemical studies of adult and fetal human testes and testicular germ-cell tumors. ALP reacting with all six antibodies was defined as placental, whereas ALP reacting with some but not all antibodies was labeled as placental-like. ALP reacting with one of the monoclonal antibodies that recognizes a determinant common to intestinal and placental ALP was tentatively considered probably intestinal, unless it reacted with any other monoclonal placental specific antibody. Using this approach, the authors have identified placental ALP in 4 of 7 seminomas, 3 of 7 tumors composed in part or fully of embryonal carcinoma, and 1 yolk sac carcinoma. Placental-like ALP was identified in 2 additional seminomas and 4 embryonal carcinoma-containing tumors, whereas 1 seminoma and 1 benign teratoma were devoid of either placental or placental-like ALP. Trophoblastic giant cells in 2 seminomas and 3 teratocarcinomas expressed only the antigenic determinant common to placental and intestinal ALP. The authors thus show that testicular tumor cells may express either placental or placental-like ALP and that in some instances, the tumor isozyme is antigenically different from ALP found on either fetal or adult testicular germ cells.