Relationship between semi-quantitative thyroid palpation and total thyroxine concentration in cats with and without hyperthyroidism.

In 155 cats, both with and without clinical signs of hyperthyroidism, total thyroxine (TT4) concentrations were compared to a sensitive, semi-quantitative thyroid palpation technique. On the basis of TT4 concentrations, 23 of the 155 cats were classified as hyperthyroid. The size of individual thyroid glands was scored between '0' (non-palpable) and a maximum of '6'. One or more enlarged thyroid glands (score >0) were palpated in 22 of the 23 hyperthyroid cats and in 78 of the 132 euthyroid cats. However, none of the 132 euthyroid cats had a thyroid lobe score of greater than '3' whereas 18 of the 23 hyperthyroid cats had a thyroid lobe score of '4' or greater, and in two of the five that had scores below '4' there was evidence of intrathoracic functional thyroid tissue on scintigraphy.

Palpable thyroid and parathyroid nodules in asymptomatic cats.

Client-owned cats underwent surgery to remove palpable cervical masses in cats with normal total T4 values and no clinical signs of hyperthyroidism, renal disease, or hyperparathyroidism. Non-functional thyroid and parathyroid adenomas were found and identified by histopathological examination. The significance of these findings is discussed in relation to palpating a goiterous mass in an asymptomatic cat.

An advanced digital image-capture computer system for gross specimens: a substitute for gross description.

The description of macroscopic appearances of surgically excised specimens together with the sites of specimen sampling form an important component in the documentation of a histopathology specimen. Unfortunately, accuracy of description depends on the vocabulary and descriptive prowess of the pathologist which means that the result can be variable. Transcription of the dictated word also takes time and involves typists. We developed a user-friendly, high-resolution image capture system that will supplement word description of macroscopic specimens and has the potential to replace it completely. It also provides the opportunity of revisiting traditional methods of documenting specimens with words, allowing the production of more relevant and user-friendly reports containing information relevant to clinical management and supplemented by high-resolution digital images. The accompanying Windows-based software has capabilities of generating the entire histopathology report and allows rapid retrospective searches through any one of several common search parameters. The stored images form a powerful database for teaching and research. Connection to remote transmission facilities allows teleconferencing and telepathology consultations.

Microwave-stimulated Jones-Marres method for staining fungi in brain tissue of immunocompromised patients.

Life-threatening fungal infections have increased significantly in the past decade due to the rising number of immunocompromised patients. Serological diagnosis of most fungal infections is unreliable and blood cultures are positive in only 50% of premortem cases; therefore, tissue sampling together with fast, reliable staining of fungi should be carried out to reach the correct, timely diagnosis. We developed, partly serendipitously, a microwave silver staining method for fungi in histological sections. During a differentiation step in periodic acid, background staining is removed. This rapid staining method can be combined with immunostaining, for example, the alpha smooth muscle actin method, to visualize blood vessels. Silver staining results were optimized using the recently developed MicroMED BASIC microwave labstation for histology (Milestone srl, Italy), featuring no-touch temperature measurements and PC control.

Visualization of vaginal flora in cervical smears using a modified microwave silver-staining method.

The advantage of studying the vaginal flora to determine the bacteria and fungi present in cervical smears (as opposed to cultivation of these micro-organisms) is that the micro-organisms can be observed in their natural habitat. However, they are only faintly stained by the conventional Papanicolaou method. Accordingly, contrast is weak and visualization poor. For this reason, we developed a modified microwave silver-staining method that can be performed retrospectively on stained smears. Bacteria and fungi stain distinctly black and can be studied in greater detail, and their inter-relationship can be visualized. Haematoxylin or Eosin counterstain allows us to visualize vaginal inhabitants in relation to epithelial cells. In the series presented here, we show that a modified microwave silver-staining method is well suited to studying the ecology of micro-organisms in smears taken from women presenting to their doctor with clinical symptoms. Using this staining method, we have shown that lactobacilli overgrowth is associated with symptoms.

Multiple piloleiomyomas in a cat.

The microscopical and immunohistochemical features of a rare cutaneous leiomyoma in a cat are described. The principal characteristics of this tumour were multiple bundles of smooth muscle, either discrete or conjoined, with numerous collagen fibres between the muscle cells, and osteoid metaplasia. The tumour was designated a piloleiomyoma as it was considered to have arisen from the arrector pili muscles.

A comparative study of cell proliferation markers in breast carcinomas.

Aims-To investigate the tumour cell proliferative index obtained by immunostaining of paraffin wax sections of 30 cases of breast carcinoma with monoclonal antibodies MIB1, KiS1 and KiS5, and polyclonal Ki67 antisera to the Ki67 antigen and 19A2 and PC10 antibodies to proliferating cell nuclear antigen and the possible correlation between these indices and that of monoclonal Ki67 antibody in frozen sections of the same tumours.Methods-All tumour samples had been uniformly fixed and processed and sections were subjected to microwave antigen retrieval before immunostaining in all instances except for monoclonal Ki67 antibody which was used in cryostat sections. Tumour cell proliferative indices were evaluated by two independent examiners, each counting 500 tumour cells with the aid of a cross-hatched grid.Results-Proliferative indices obtained with MIB1, polyclonal Ki67, KiS1, and KiS5 correlated with those obtained with monoclonal Ki67 in frozen sections. Proliferative indices obtained with monoclonal 19A2 and PC10 showed no correlation with those of monoclonal Ki67 antibody. The staining obtained with MIB1 was the most intense and the easiest to read.Conclusions-Monoclonal antibodies MIB1, KiS1 and KiS5 and polyclonal Ki67 antiserum appear to be suitable substitutes for monoclonal antibody Ki67 in the assessment of tumour cell proliferative index. As these reagents are all immunoreactive in paraffin wax sections, they overcome the requirement for frozen tissue for immunostaining with monoclonal Ki67.

Dual staining of lymphocyte membrane antigens with colloidal gold and biotinylated horseradish peroxidase.

Double immunoenzymatic labelling procedures for the localization of antigens on cells in tissue sections using horseradish peroxidase (HRP) and alkaline phosphatase have been described previously, but mainly for detecting antigens on different cells. With this type of staining when two antigens are present on the same cell, an optimal colour combination that shows a high contrast between the basic colour of each enzyme substrate product is difficult to achieve and the interpretation of their mixed colour intermediate is subjective. We present a method for the simultaneous demonstration of two antigens on the same cell. The method can be used to label either single cells in suspension, or cells in paraffin fixed tissue, using a combination of a particulate label, colloidal immunogold-silver, and an enzymatic label HRP-DAB. The method is easy to perform and utilises commercially available staining kits.

Bacillus Calmette-Guérin-treated superficial bladder cancer: correlation of morphology with immunophenotyping.

Thirty-five biopsies from 19 patients with superficial transitional cell carcinoma of the bladder, treated with intravesical bacillus Calmette-Guérin (BCG), were assessed histologically and immunohistochemically. Pretreatment biopsies were available for comparison in all cases and five cases of non-specific cystitis were also examined. The inflammatory infiltrate was assessed with a streptavidin-biotin-peroxidase method using UCHL1, MT1, LN3, L26, HAM56, MAC387, Leu7 and anti-S100 in paraffin sections, and in 18 specimens were frozen tissues were available, Leu1, 2, 3, 4, 14, OKT10, HLA-DR and anti-Tac antibodies were applied. Post-treatment bladder biopsies showed severe oedema and a variable degree of inflammation. A granulomatous inflammation was seen in 11 cases, with granulomas present in six prostatic biopsies and acid-fast bacilli in two cases. The lymphoid infiltrate in all biopsies were largely T lymphocytes with a predominance of T helper cells present, often as a band-like infiltrate pressing against the residual epithelium, or the denuded bladder surface, and distributed in the vicinity of the granulomas. Activated lymphocytes were prominent in seven cases, although a moderate infiltrate of such cells was seen in all instances. Tac antigen was only occasionally expressed, and in a few NK cells were present among the infiltrates. In eight cases, HLA-DR was expressed in epithelial cells following BCG treatment, whereas all pre-treatment epithelial were negative.

Accelerated immunohistochemical staining by microwaves.

The application of microwave irradiation is extended to include the acceleration of the streptavidin-biotin peroxidase staining procedure for the detection of a wide range of antigens in paraffin-embedded sections of both normal and neoplastic tissues. Microwave irradiation is used for all procedural steps requiring incubation so that the entire staining procedure can be completed within 20 min. The technique is simple and convenient, with uniform irradiation times for different tissue antigens. The results obtained equalled that of conventional staining procedures.

"Thick" cell membranes revealed by immunocytochemical staining: a clue to the diagnosis of mesothelioma.

The distinction of malignant mesothelioma from metastatic adenocarcinoma in pleural effusions and biopsies is frequently a diagnostic problem. Immunocytochemical staining of 13 malignant mesotheliomas, eight primary adenocarcinomas of the lung, five metastatic adenocarcinomas of the lung, and 20 primary adenocarcinomas in extrapulmonary sites with a monoclonal antibody to epithelial membrane antigen (EMA) revealed "thick" cell membranes in all cases of mesothelioma. This distinctive pattern of staining was seen in the periphery of cell clusters and circumferentially around individual cells in cytologic preparations, cell blocks, and tissue sections. Intracellular and intercellular acini were also outlined by anti-EMA, and long intraluminal microvillous projections were demonstrated. Weak cytoplasmic staining was only rarely seen in mesothelioma cells. This membranous staining pattern was not observed in adenocarcinomas, which displayed strong and diffuse cytoplasmic staining. The immunocytochemical demonstration of thick and spiky membranes circumferentially disposed around individual cells corresponds to aberrant microvilli, a diagnostic clue in the recognition of malignant mesothelioma.

Antigen preservation in microwave-irradiated tissues: a comparison with formaldehyde fixation.

A wide range of immunoreactive antigens commonly investigated in diagnostic pathology were evaluated in tissues subjected to fixation by microwave irradiation in normal saline to 63 degrees C. All of the 23 antigens studied were well preserved and compared with corresponding tissues conventionally fixed by formaldehyde for periods of 16 and 18 h, respectively, immunostaining in microwave-irradiated tissues was clearly superior, being more intense and also more extensive. Proteolytic digestion was not necessary for irradiated tissues except for the staining of cytokeratins and desmin. It was also possible to stain for Leu-14 (CD22) and Leu-1 (CD5), whereas these lymphocyte antigens were lost during formaldehyde fixation. Cytomorphological details were well preserved in the rapidly-fixed irradiated tissues.

Cytokeratin and vimentin intermediate filament proteins in benign and neoplastic prostatic epithelium.

Prostatic samples from 30 hyperplastic prostates and 61 prostatic adenocarcinomas were examined for vimentin and cytokeratins. The co-expression of cytokeratins and vimentin was found in all benign prostatic epithelium and in 83% of adenocarcinomas. Benign prostatic epithelium showed vimentin intermediate filaments distinctively distributed in the basal regions and as paranuclear sheaves along the long axis of the cell. This pattern of vimentin staining was seen in adenocarcinomas with low Gleason scores, whereas high-grade tumours showed intense diffuse perinuclear staining.

Growth fractions in human prostatic carcinoma determined by Ki-67 immunostaining.

Until recently, [3H]-thymidine incorporation, DNA analysis by flow cytometry, and cell doubling times have been the main methods of studying tumour cell kinetics. All these techniques are laborious, expensive, and difficult to perform in a routine diagnostic laboratory. This study examined fresh frozen sections from 31 prostatic biopsy specimens with the hybridoma antibody Ki-67, a marker of proliferating cells, using a modified avidin-biotin-peroxidase complex technique. The percentage of glandular cells decorated by this antibody, representing the growth fraction, was determined for both benign and malignant samples. Benign prostatic glands showed an average Ki-67 score of 4 per cent, significantly less than the 16.3 per cent mean growth fraction found in prostatic carcinomas. There was a significant correlation between the tumour growth fraction as assessed by Ki-67 staining, and the histological grade. A positive correlation was also found between the Ki-67 score and the intensity of staining, and a definite trend was noted between the Ki-67 score and the tumour clinical stage. Ki-67 promises to be a useful marker in determining the prognosis of prostatic cancer.

The application of the avidin-biotin technique to previously stained histological sections.

Using the sensitive avidin-biotin peroxidase technique, a wide variety of tissue antigens can be detected in standard histological sections of both normal and pathological tissues previously stained with hematoxylin and eosin. There appears to be no detectable reduction of sensitivity with this method of "restaining" compared to the standard immunoperoxidase procedure applied to unstained tissue sections. This technique makes it possible for retrospective identification of tissue antigens when insufficient unstained material is available.

Atypical fibroxanthoma of the skin: a clinicopathological and immunohistochemical study and a discussion of its histogenesis.

The morphological and immunohistochemical characteristics of 37 atypical fibroxanthomas of the skin were examined. Twenty-four tumours were nodular ulcerative lesions on the head and face of patients with a median age of 75 years, whereas 13 tumours occurred on the trunk and limbs of patients with a median age of 48 years. Both pleomorphic polygonal and giant cells as well as the spindle cell component of the tumours stained for the histiocytic markers alpha 1-antichymotrypsin, alpha 1-antitrypsin, lysozyme and, less frequently, for ferritin. Leu M1 antigen and peanut agglutinin receptors were not demonstrable in tumour cells. This antigenic profile was contrasted with the findings in six cases of dermatofibroma which were largely not reactive with the antisera used. The immunohistochemical findings in atypical fibroxanthomas suggest that they represent a homogeneous group of tumours which are related to tissue histiocytes. These results are discussed in the context of the published findings in other so-called fibrohistiocytic tumours including dermatofibrosarcoma protuberans and malignant fibrous histiocytoma. The diagnoses in three cases coded as atypical fibroxanthomas were revised on the basis of their showing a different immunohistochemical profile.

Criteria for the diagnosis of primary endocrine carcinoma of the skin (Merkel cell carcinoma). A histological, immunohistochemical and ultrastructural study of 13 cases.

Thirteen cases of primary endocrine carcinoma of the skin (Merkel cell carcinoma) were reviewed with the aim of defining the morphological, immunohistochemical and ultrastructural criteria for diagnosis. The tumour cells were characterized by their scanty cytoplasm, generally small uniform nuclei with finely dispersed chromatin and multiple small nucleoli. Nuclear shapes varied from round to spindle, with larger and pleomorphic forms predominating in 2 tumours. A striking feature seen in 12 tumours was the occurrence of a "ball-in-mitt" pattern represented by 1 or 2 crescentic tumour cells closely wrapped around an oval cell. Staining for neuron-specific enolase was the most consistent marker of the tumour and the characteristic juxtanuclear globular staining for keratin and cytokeratin and the occasional coexpression of neurofilament set this tumour apart from other cutaneous neoplasms, in particular, metastatic carcinoid tumours and oat cell carcinoma from the lung. The fine structural features of note were striking paranuclear or juxtanuclear whorls of intermediate filaments, seen in 7 cases, the presence of variable numbers of membrane-bound dense core granules of 80-150 nm diameter in all cases and cytoplasmic spinous or microvillous projections containing microfilaments in 4 cases. Less consistent characteristics of primary endocrine carcinomas of the skin included cell moulding, argyrophilia and immunohistochemical staining for ACTH, VIP and calcitonin. The high frequency of vessel invasion in this series is in keeping with the high rate of local recurrence, lymph node metastases and visceral dissemination reported. The distinction from other similar appearing tumours in the skin is discussed.

Rapid immunoperoxidase staining of lymphocyte antigens using microwave irradiation.

The innovative application of microwave irradiation in the immunoperoxidase staining of a wide range of labile lymphocyte antigens is described. Fifteen second irradiation at 320 watts produced excellent adherence of the frozen sections to the glass slides without loss of surface antigen staining. This brief procedure eliminated the need for much longer periods of freeze drying or drying at 4 degrees C. Microwave irradiation was also used to accelerate the incubation of the primary antibody. Thirty seconds of irradiation at 320 watts produced specific antigen staining of an intensity which was as good as that obtained with much longer durations of incubations at room temperature. Cytomorphologic preservation was also good and background staining was minimal.