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Biomedical subjects

M E Boon

Publications and source records attributed to M E Boon.

At least 163 records · Page 9Linked to original sources
PubMed1981

Learning from quantitation.

An important question in the effective use of quantitation in cytology is how to make the link between mathematical parameters and the cytologist. As an example, an investigation is discussed in which cell images from benign mesothelial proliferations were distinguished by quantitative characteristics from those of malignant mesotheliomas originating in the pleura and peritoneum. The significant differences between the three groups in nuclear size, cytoplasmic size and nuclear-cytoplasmic (N/C) ratio could partly be explained by the vacuolation patterns of the exfoliated mesothelial cells in each. Moreover, the primary site of the tumor could be predicted by the quantitative cell parameters. The characteristics of cytologic images, once known to be important, are easily incorporated into standard diagnosis. Thus it now seems possible for the cytologist to differentiate between the three groups by using the cytomorphologic features that relate to the computed parameters. This shows the value of translating computerized parameters into cytomorphologic criteria.

Ascitic Fluid↗
PubMed1980

DNA content of normal and neoplastic urothelial cells. A comparative cytofluorometric and cytomorphologic study.

In search for suitable parameters to detect neoplastic urothelial cells in Acriflavine-Feulgen-SITS stained specimen we compared the cytofluorometric DNA content with the morphology of normal urothelial cells (bladder scrapings) and neoplastic urothelial cells from grade 1, 2, 3 and 4 tumors. An individual normal urothelial cell could not be distinguished from a grade 1 tumor cell, neither morphologically nor fluorometrically. However, the shape of the histograms of DNA measurements of the cell populations of respectively normal bladder scrapings and grade 1 tumors differs. It is postulated that also morphometry of these cell populations may be of some aid to distinguish well-differentiated neoplastic cells from normal urothelial cells. Seventy-one percent of the morphologically malignant cells in the grade 2,3 and 4 tumor samples could be identified by applying the combined parameters: high DNA content (greater than 5 C) and nuclear-cytoplasmic ratio (greater than 0.5) and all grade 2, 3 and 4 tumor samples contained cells which were objectively classified as malignant. Using the same parameters morphologically malignant cells could be distinguished from normal, polyploid umbrella cells, thus these malignant cells are detectable objectively without using chromatin pattern as parameter.

Cell Nucleus↗
PubMedDate not supplied

The predictive value of immature dysplastic cells in CIN I and II smears.

This paper reports the predictive value of a subclassification of cervical intraepithelial neoplasm grade I and II smears. The follow-up period was a maximum of 1 yr. The subclassification was based on the presence of immature dysplastic cells ("IDC present") in the original smear. In 267 cases, these immature dysplastic cells were morphometrically defined. In an additional group of 14 cases, the immature dysplastic cells were exclusively in dense cell groupings which were unsuitable for morphometrical analysis; thus, only visual classification was possible. The predictive value negative (follow-up with negative findings or CIN I or II) of this method was 1.00. The predictive value positive (follow-up with CIN III) was 0.14. The sensitivity was 0.98 and the specificity 0.57. On the basis of this subclassification, cases that should be directly referred for further investigation can be selected. It is justifiable to follow-up the cases that are "IDC absent" with cytology alone. The described method is relatively simple, and can be performed in routine diagnostic laboratories. It is very valuable in settings where most of the initial smears and the follow-up are in the hands of general practitioners. Using the described method, patients who should be referred to the gynecologist for colposcopy and histology can be selected.

Cervix Uteri↗
PubMedDate not supplied

Formaldehyde fixation and microwave irradiation.

Formaldehyde is the most commonly used fixative in pathology laboratories. However, due to time pressures, this fixative is often not optimally exploited. The majority of biopsies are only partly fixed when histoprocessing is started, with adverse effects. This paper reports how formaldehyde fixation is improved, by using 1.5 min of microwave irradiation of tissue previously soaked for four hours in the fixation solution. It is argued that this beneficial effect of microwave irradiation can be attributed to the acceleration of the reaction of formaldehyde to the tissue. Formation of free formaldehyde, by the dehydration of methylene glycol present in the tissue when the irradiation starts, is also enhanced. Five different formaldehyde-containing fixatives were evaluated, using five different working protocols. Spleen was taken as a suitable tissue for these tests. The technique described leads to uniform microscopical results. It is a simple method and is suitable for use in routine laboratories.

Formaldehyde↗
PubMedDate not supplied

Histoprocessing with the microwave oven: an update.

This paper evaluates and extends the novel method of preparing tissue blocks for paraffin sections within 30 to 60 min, that was proposed in early 1985 in a paper by Boon et al. (1986). More than 2 years' additional experience and testing various microwave ovens has led to new protocols reported in this paper. Results are given of testing (i) an especially designed microwave oven for histoprocessing, (ii) microwavable reagents, (iii) processing larger numbers of specimens simultaneously, (iv) handling different types and sizes of tissue. It is concluded that effective temperature control offers substantial advantages. In addition, the possibilities of performing routine diagnostic pathology omitting formalin altogether are sketched.

Female↗
PubMedDate not supplied

Understanding microwave-stimulated Romanowsky--Giemsa staining of plastic embedded bone marrow.

Bone marrow smears were made and fixed in methanol or formaldehyde. Marrow sections of various thicknesses were also prepared from formaldehyde fixed marrows embedded in paraffin or plastic (glycol methacrylate). The different smears and sections were then stained by a Romanowsky--Giemsa procedure. Some specimens were stained using a standard microwave-stimulated method previously used diagnostically. The effects of technical variations were studied, including degree of microwave irradiation and the staining time. Comparisons of the resulting staining outcomes showed that microwave stimulated Romanowsky--Giemsa staining of plastic sections is a rate controlled process. Unusual aspects of the staining pattern of plastic sections (namely the purple basophilic cytoplasms and nucleoli, and blue chromatin) are due to microwave stimulation and formaldehyde fixation respectively.

Animals↗
PubMedDate not supplied

Microwave-aided technique to detect bromodeoxyuridine in S-phase cells using immunogold-silver staining and plastic-embedded sections.

A technique is described to detect bromodeoxyuridine (BrdU) incorporated by cells in S-phase, with a monoclonal antibody, using removable plastic embedding and immunogold-silver staining (IGSS). The incubation times were reduced and the immunological reactions enhanced by microwave irradiation. The embedding in methyl methacrylate enabled us to make thinner sections and it improved the quality of the preparations. The methyl methacrylate did not hinder the reaction of BrdU with the antibody because it could be removed prior to the IGSS procedure. The IGSS procedure appeared to be very sensitive, requiring lower concentrations of the antibodies than other methods. The use of microwave irradiation shortened the time needed to stain a section from 7 to less than 4 h. Furthermore, using microwave irradiation, the concentration of the antibodies needed could be reduced even further compared with the normal IGSS procedure. In sections of the mouse testis and small intestine only nuclei of cells known to be able to proliferate appeared BrdU positive. The non-specific background staining was found to be negligible. In testes of mice that received both 3H-thymidine and BrdU more than 95% of the radioactively labelled cells also showed BrdU label and vice versa. This indicates that both methods are equally sensitive for detecting cells in S-phase.

Animals↗
PubMedDate not supplied

Preservation of structure and cytochemical reactivity at the ultrastructural level, using microwave irradiation.

The application of microwave irradiation at various steps in 'normal' EM procedures has been investigated. 1. The temperatures in various aqueous volumes were measured during microwave irradiation. Increases were small and the final temperature could be controlled by cooling the glass base plate. 2. The influence of microwave irradiation on the various fixation schedules for the electron microscopy of single cell populations was studied and the results favoured the idea that by the application of microwave irradiation a more life-like ultrastructural preservation could be obtained. 3. Peroxidase-like activity in erythrocytes, acid phosphatase activity in resident macrophages and peroxidase activity in monocyte granules was apparently not influenced by microwave irradiation during aldehyde fixation and incubation. 4. The anticipated microwave-induced penetration enhancement of cerium ions in a cytochemical reaction procedure to detect acid phosphatase activity in resident macrophages was not observed.

Acid Phosphatase↗
PubMedDate not supplied

Microwave stimulation of an immunological reaction (CEA/anti-CEA) and its use in immunohistochemistry.

The effects of microwave stimulation on different aspects of immunohistochemical reactions were studied using Elisa as a model system. This technique allows monitoring of (i) the rates and recoveries of different antigen-antibody reactions, (ii) the formation of avidin/biotin complexes, (iii) the prevention of non-specific activity and (iv) the washing procedures. The binding reactions seemed to follow second-order kinetics, and reaction rates were considerably enhanced by microwave stimulation. The steps were aimed at preventing non-specific reactivity and the washing procedures could all be shortened to seconds. Such rate increases are far too large to be explained solely by the modest increase in temperature. Furthermore, microwave irradiation caused a major reduction in the yield of antigen-antibody complexes. This inhibitory effect can be compensated for by increasing the concentration of antibody. No significant effects of the microwaves on the antigen-antibody complexes were found once the complexes had been formed. The obtained results were then applied to the demonstration of carcinoembryonic antigen in histological material. By employing this method a complete immunohistochemical staining of a hydrated tissue section could be performed within 15 min, excluding the final development of colours.

Antigen-Antibody Complex↗
PubMedDate not supplied

Brain enzyme histochemistry following stabilization by microwave irradiation.

The activities of various enzymes present in brain homogenates were assayed biochemically (a) with no pretreatment, (b) following a standard microwave treatment in saline and (c) after a standard microwave treatment in formalin. All enzyme activity was lost after the microwave - formalin in treatment. Following microwave - saline treatment, the activities of alkaline phosphatase, 5'-nucleotidase, isocitrate and succinate dehydrogenases were reduced. In contrast, the activities of lactate and malate dehydrogenases were unchanged, and that of acetylcholinesterase apparently increased. Analogous outcomes were seen following attempted histochemical demonstrations of these enzymes. Thus satisfactory histochemical demonstration of all enzymes was achieved (except with alkaline phosphatase, lactate and malate dehydrogenases) following the microwave-saline pretreatment. Since acid phosphatase, catalase and peroxidase were also successfully demonstrated, it seems that microwave-saline pretreatments permit both retention of sufficient enzyme activity for histochemical demonstration to occur and retention of sufficient structural integrity for critical morphological investigations. Since the failure to stain the sites of lactate and malate dehydrogenases is not due to microwave inactivation of these enzymes, their demonstration may be possible by varying the staining procedures.

Animals↗
PubMedDate not supplied

Microwave irradiation in label-detection for diagnostic DNA-in situ hybridization.

A DNA-in situ hybridization protocol was adapted for application to sections of routinely processed paraffin embedded material. This protocol was developed previously for detecting DNA-virus infected cells in whole cell preparations and employs biotinylated DNA as probe. Three different biotin detection methods were optimized and applied. The first uses streptavidin and a biotinylated complex of alkaline phosphatase, the second consists of an immunogold-silver staining, and the third of a peroxidase technique using a silver amplification. The alkaline phosphatase method was the most rapid, and as sensitive as the immunogold-silver staining. The peroxidase method was the most sensitive. Microwave irradiation was applied to the different incubation steps of these three detection methods. Short incubations with microwave irradiation gave results comparable to those obtained with conventional incubations, when streptavidin, antibiotin, complexed alkaline phosphatase, or gold labelled goat antirabbit were used. It was thus shown that microwave irradiation creates the possibility of a very rapid label-detection for nonradioactive DNA-in situ hybridization.

Alkaline Phosphatase↗
PubMedDate not supplied

Microwave stabilization versus chemical fixation. A morphometric study in glycolmethacrylate- and paraffin-embedded tissue.

A morphological and morphometrical study was performed on testicular cells after microwave stabilization of the tissue while immersed in phosphate buffered saline (PBS), 0.9 NaCl or Tris-HCl. Fixation in Carnoy's fluid without irradiation was chosen as a control chemical fixation method. After microwave stabilization or chemical fixation, the testes were embedded in paraffin or in plastic (glycolmethacrylate). An excellent morphology, comparable to that after chemical fixation in Carnoy's fluid, was observed in the plastic sections of tissue irradiated in PBS or NaCl, even when the sections were subsequently treated with an aggressive reagent at high temperature, required for the Feulgen reaction. The nuclear area of the microwave-stabilized Sertoli cells was 37-46% smaller in haematoxylin-eosin stained, paraffin sections in comparison with that in the glycolmethacrylate sections. The microwave-stabilized, paraffin-embedded tissue was much more vulnerable to the hot HCl treatment of the Feulgen staining than the chemically fixed tissue, resulting in an additional 10-20% decrease in nuclear size. The latter finding is particularly important for quantitative microscopy, where the Feulgen staining method is often employed.

Animals↗
PubMedDate not supplied

Notes on the application of microwaves in histopathology.

In this article two applications of microwaves in histopathology, microwave-stimulated staining of tissue sections and microwave-stimulated fixation of cryostat sections, are reviewed. For a good understanding of the influence of microwaves on physico-chemical processes like staining and fixation the relevant physics are included. Major advantages of microwave techniques are speed and/or improved quality. The cryostat-microwave technique appears to be well-suited for the demonstration of intermediate filament proteins: the sensitivity of monoclonal antibodies directed against keratins and vimentin can be substantially increased using the ethanol based fixative Kryofix.

Histocytochemistry↗