Microwaves for chromogenic in situ hybridization.

In situ hybridization can be employed in formalin-fixed, paraffin-embedded tissue sections (FFPT) and allows direct visualization of amplified genes and chromosomes in individual cell nuclei. Fluorescence in situ hybridization (FISH) is the most widely employed method, but the fluorescence preparations suffer from the main disadvantages of fading over time and poor visualization, the latter making it difficult to accurately separate invasive from in situ cancer cells. Chromogenic in situ hybridization (CISH) is a viable alternative to FISH in FFPT as it employs a peroxidase reaction to visualize the chromogen thus allowing the convenience of bright field microscopy and the correlation of the visualized gene amplification with cytomorphology.

Standardization in immunohistology.

The rapid acceptance of immunohistology as an invaluable adjunct to morphologic diagnosis has been possible because of the development of new and more sensitive antibodies and detection systems that allow its application to formalin-fixed, paraffin-embedded tissue (FFPT). More importantly, antigen-retrieval techniques have resulted in some degree of consistency allowing immunohistology to be used reliably as a diagnostic tool. The advent of prognostic and predictive biomarkers, and the desire for individualized therapy has resulted in mounting pressure to employ the immunohistological assay in a quantitative manner.

Glomeruloid peritoneal implants in ovarian serous borderline tumours--distinction between invasive and non-invasive implants and pathogenesis.

Aims: To determine whether or not the glomeruloid implants (GI) composed of papillary cores within clear spaces lined by mesothelial cells or tumour cells located in superficial or deep peritoneal tissue in ovarian serous borderline tumours (SBTs) are invasive.

Methods And Results: We examined the differences in incidence, histological and immunohistochemical findings among three groups: 100 GI with mesothelial cells lining clear space (type I), 100 GI with tumour cells lining clear space (type II), and 100 invasive implants with clefts but no lining cells from 30 cases of SBT with peritoneal implants. The type I lesion had characteristics of non-invasive implants with a tendency for smooth contours (100/100), superficial location (71/100), absence of desmoplasia (100/100) and absence of surrounding destructive invasion (100/100), In contrast, type II GI had irregular contours (67/100), deep location (93/100), presence of desmoplastic reaction (100/100) and presence of destructive invasion (12/100).

p16, cyclin D1, Ki-67, and AMACR as markers for dysplasia in Barrett esophagus.

Barrett esophagus (BE) is an established precursor of esophageal adenocarcinoma (AdenoCa). One hundred and one cases of BE diagnosed by esophageal biopsy and resections were examined morphologically for dysplasia. These were categorized as BE without dysplasia (n=25), indefinite for dysplasia (IND, n=17), low-grade dysplasia (LGD, n=18), high-grade dysplasia (HGD, n=15), and AdenoCa (n=26).

Epidemiology and carcinogenesis of hepatocellular carcinoma.

The incidence of hepatocellular carcinoma (HCC) shows marked variation worldwide but the magnitude of this tumor is reflected by the occurrence of at least 1 million new cases annually and the uniformly dismal outlook with median survivals of <25 months after resection and <6 months with symptomatic treatment. The strikingly uneven distribution of this tumor parallels the prevalence of hepatitis B infection with rising incidence in western countries attributed to hepatitis C infection. Chronic hepatitis and cirrhosis constitute the major preneoplastic conditions in the majority of HCCs and may be related to other etiologic agents such as environmental chemical carcinogens including nitrites, hydrocarbons, solvents, organochlorine pesticides, and the chemicals in processed foods, cleaning agents, cosmetics and pharmaceuticals, as well as plant toxins such as anatoxins produced by fungi that cause spoilage of grain and food in the tropics.

Vascular and myometrial changes in the human uterus at term.

At term, amniotic fluid contents may mediate the onset of labor through the activation of amniotic fluid macrophages and their migration into the myometrium. To test this concept, the authors examine the histological changes that occur in myometrial biopsies at term prior to (n = 53) and during (n = 15) labor. Biopsies were stained with an antimacrophage antibody, anti-CD34 (endothelial cells), and anti-caspase 3 (apoptotic cells).

The pathology of dengue hemorrhagic fever.

An estimated 2.5 billion people are at risk of dengue infection, and of the 100 million cases of dengue fever per year, up to 500,000 develop dengue hemorrhagic fever (DHF) or dengue shock syndrome (DSS), the life-threatening forms of the infection. The large majority of DHF/DSS occurs as the result of a secondary infection with a different serotype of the virus.

Microwave enhancement of CISH for HER2 oncogene.

We describe a modification to the prescribed procedure for the Zymed Spot-Light HER2 chromogenic in situ hybridization kit (84-0146, Zymed Laboratories, San Francisco, CA) by substituting the heat pretreatment step with MW irradiation in citrate buffer 10 mmol/L at pH 6.0 at 98 degrees C for 10 minutes and repeating the procedure afterenzyme digestion with time and temperature controlled in the Mega T/ T oven (Milestone s.r.

How does antigen retrieval work?

The introduction of antigen retrieval has enabled immunohistology to become an integral component of morphologic diagnosis, routinely employed in cancer diagnosis, and for the identification of therapeutic and prognostic markers. The mechanism of antigen retrieval, however, remains speculative with the key to our understanding embedded in the actions of formaldehyde on proteins. One commonly held concept is that heat primarily breaks down protein cross-linkages that occur with aldehyde fixation, thus "unmasking" protein epitopes of interest.

The morphological spectrum of lymphadenopathy in HIV infected patients.

Aims: To study the histological spectrum of lymphadenopathy in human immunodeficiency virus (HIV) infected Thai patients.

Methods: Lymph nodes from 55 HIV infected patients were accessioned over a 19 month period in two pathology laboratories in Bangkok, Thailand. These were examined with H&E, Ziehl-Neelsen, periodic acid-Schiff (PAS), PAS with diastase (PAS/D), Gram and methenamine stains.

Diagnostic 'errors' in anatomical pathology: relevance to Australian laboratories.

Failure to recognise that anatomical pathology diagnosis is a process of cognitive interpretation of the morphological features present in a small tissue sample has led to the public misperception that the process is infallible. The absence of a universally accepted definition of diagnostic error makes comparison of error rates impossible and one large study of laboratories in the United States shows a significant error rate of about 5%, most of which have no major impact on patient management. A recent review of the work of one pathologist in New South Wales confirms a lack of appreciation in medical administration that variable diagnostic thresholds result in an inherent fallibility of anatomical pathology diagnoses.

Refinement of immunohistologic parameters for Her2/neu scoring validation by FISH and CISH.

The conventional method of scoring Her2/neu immunostaining is recognized to result in a high false-positive rate among 2+ cases when compared with results obtained with fluorescence in situ hybridization (FISH); however, costs and convenience dictates that immunohistochemistry remains the screening test for Her2/neu status in patients with breast cancer. We describe refined criteria for scoring of Her2/neu on the basis of anatomic localization rather than the subjective assessment of intensity. The presence of a circumferential tram track pattern that results from the staining of apposing cell membranes in >25% of the tumor cells was necessary for a 3+ score (Her2/neu overexpressed) and the presence of the tram track pattern in <25% was scored 2+ (equivocal); granular and fragmented membrane staining was scored 1+ (negative).

Controversies in the assessment of HER-2: more questions than answers.

Human epidermal growth factor receptor 2 (HER-2) is over-expressed in 15% to 30% of breast cancers and is a poor prognostic marker in node-positive patients. HER-2 expression is an indicator of greater sensitivity to anthracycline-based chemotherapy and is the major criterion for selection for treatment with the anti-HER-2 antibody trastuzumab (Herceptin). Fluorescence in situ hybridization and immunohistochemistry (IHC) are the 2 most commonly used methods for detection of the gene and protein, respectively.

Cytokeratin 20 and Ki-67 to distinguish carcinoma in situ from flat non-neoplastic urothelium.

Urothelial carcinoma in situ (CIS) is a high-grade neoplasm and an indicator of recurrence and progression that requires specific treatment. The distinction of CIS from flat non-neoplastic urothelium, in particular dysplasia, on the basis of histologic features is often difficult, and this study aims to validate cytokeratin 20 (CK20) and Ki-67 as discriminatory markers for this purpose. Immunostaining of these markers was applied to 26 cases of CIS, 14 atypia of unknown significance, 4 dysplasia, 6 normal, and 9 hyperplastic urothelium.

Immunohistological localisation of human FAT1 (hFAT) protein in 326 breast cancers. Does this adhesion molecule have a role in pathogenesis?

Aims: To examine the immunohistological expression in human breast cancers of human FAT1 (hFAT) protein, a recently described member of the cadherin superfamily, and its correlation with histological type and grade.

Methods: A total of 326 cases of invasive and in situ breast cancer representing a broad spectrum of histological subtypes were immunostained with affinity-purified rabbit antibodies produced to the cytoplasmic region of hFAT using a standard avidin-biotin system. Staining intensity was arbitrarily graded on a scale of 0 to 3.

Calretinin, CD34, and alpha-smooth muscle actin in the identification of peritoneal invasive implants of serous borderline tumors of the ovary.

The correct identification of invasive implants in the peritoneum in serous borderline tumors (SBTs) of the ovary is an important determinant of diagnosis, treatment, and prognosis. Although the histologic criteria to distinguish noninvasive from invasive implants have been defined, the distinction can still be difficult. We examined the presence and distribution of mesothelial cells, stromal fibrocytes, and myofibroblasts in invasive and noninvasive peritoneal implants in 100 noninvasive, 100 invasive, and 100 metastatic nests/foci from 20 cases of SBTs with peritoneal implants, 10 serous carcinomas with peritoneal metastasis, and 10 cases of endosalpingiosis by immunostaining for calretinin, CD34, and alpha-SMA.

Basal lamina visualization using color image processing and pattern recognition.

In histologic assessment, the absence of basal lamina is a useful feature for distinguishing invasive malignancy from benign and in situ lesions. As this feature is not possible to assess in routine H&E sections, pathologists have instead relied on histochemical and immunohistochemical stains to show components of the basal lamina such as laminin or type IV collagen. Standard image-processing software with the necessary image-processing toolbox (Matlab v5, Mathworks, Natick, MA) was used in a unique combination of color image processing and pattern recognition techniques to accentuate the collagenous stroma surrounding glands, which approximates basal lamina, in a series of benign, in situ, and invasive breast proliferations.

Multiple organ infection and the pathogenesis of SARS.

After >8,000 infections and >700 deaths worldwide, the pathogenesis of the new infectious disease, severe acute respiratory syndrome (SARS), remains poorly understood. We investigated 18 autopsies of patients who had suspected SARS; 8 cases were confirmed as SARS. We evaluated white blood cells from 22 confirmed SARS patients at various stages of the disease.