Search PubMed⌕ Search

Biomedical subjects

G R Newman

Publications and source records attributed to G R Newman.

At least 37 records · Page 2Linked to original sources
PubMed1993

The distribution of amyloid plaques in the cerebellum and brain stem in Down's syndrome and Alzheimer's disease: a light microscopical analysis.

The distribution and severity of the neuropathological changes in 57 cases of Alzheimer's disease, and 11 patients with Down's syndrome were investigated with reference to the cerebellum. A modified silver stain and a monoclonal antibody raised against amyloid beta-protein were used to identify amyloid plaques. The highest incidence of amyloid plaques in the cerebellum (93%) was found in the group of patients who developed dementia before 65 years of age. This figure dropped to 56% in those patients with dementia beginning after 75 years. In 37 of these cases the distribution of the pathological changes of the disease were also examined in the brain stem. The severity of the pathological changes in the cerebellum corresponded to the involvement of the brain stem nuclei with connections to the cerebellar cortex. The possibility that the disease process spreads to the cerebellum by involving the fibres from the brain stem is discussed with reference to previous anatomical and neurochemical studies.

Adult↗
PubMed1993

Corpora amylacea could be an indicator of neurodegeneration.

We describe an investigation of corpora amylacea (CA) in the brain tissue of Alzheimer's disease (AD) cases and normal ageing controls, using both light (LM) and electron (EM) microscopic techniques. CA populations were shown by routine histological staining of LR White resin sections with methenamine silver and PAS, and were compared with those shown by immunocytochemistry using antibodies to tau, GFAP, tubulin, ubiquitin, beta-amyloid and serum amyloid P component in serial sections. All CA were immunoreactive with anti-tau and all were unreactive with anti-beta-amyloid. Most were immunoreactive with anti-serum amyloid P component, although this was often weak in AD. CA from normal ageing brain were immunoreactive for proteins that are associated with the neuronal cytoskeleton and cell injury. CA from AD brain shared some of these but differed from those in normal ageing brain by being in much larger number and more variable in their immunoreactivity. In all CA, X-ray microanalysis illustrated the presence of the metallic elements Ca, Fe and Cu. Aluminium, often associated with AD, was not present, even in CA from AD brain. Phosphorus and sulphur, probably from phosphorylated proteins associated with degenerating cytoskeleton elements, were usually detected. In AD brain, the greater numbers of CA and their variable biochemical and elemental composition, when compared with CA in the normal ageing brain, suggests that they may derive from a number of sources both neuronal and glial as a result of the neurodegenerative disease.

Alzheimer Disease↗
PubMed1992

Dinitrophenyl (DNP) hapten sandwich staining (DHSS) procedure. A 10 year review of its principle reagents and applications.

Over the past 10 years an immunoperoxidase method using dinitrophenyl (DNP) hapten-labelled primary or secondary probes has been devised. Its widely successful application in research and diagnostic work has depended upon the development of certain key reagents. These include a novel non-deleterious DNP labelling compound, a unique multivalent monoclonal bridge antibody, and an efficient DNP hapten substituted or anti-DNP linked marker enzyme. In this article the development of these reagents and various modifications of the basic technique are reviewed in conjunction with the special applications accruing from their use.

Antibodies, Monoclonal↗
PubMed1991

Strategies for improving the cytochemical and immunocytochemical sensitivity of ultrastructurally well-preserved, resin embedded biological tissue for light and electron microscopy.

Many techniques for processing tissue into resin are available, varying from conventional room temperature to low temperature procedures. The problem is to choose an appropriate method to suit the biological specimen under study. Room temperature approaches with aldehyde and osmium fixation do not give optimal retention of immunoreactivity. Osmium can be removed from sections, but recovery of immunosensitivity is reduced. Osmium post-fixation can be omitted, but heat polymerization of resins causes tissue extraction and loss of immunoreactivity. Alternative techniques rely on the use of milder polymerization methods and avoid osmium. However, while providing an improvement, this alone is not sufficient to maximize tissue reactivity. Fixation with high concentrations of glutaraldehyde (greater than 1%) and processing into resin at either room or low temperature results in retention of similar levels of immunoreactivity. Low concentration glutaraldehyde (less than 0.2%) fixation for short periods of time (less than 60 minutes) produces improved tissue immunoreactivity and allows low concentrations of antigen at secondary sites to be detected. However, the tissue is now only minimally stabilized and is prone to extraction and conformational damage during processing. It can be partially protected by employing one of two strategies: processing at room temperature with partial dehydration (upto 70% solvent) and rapid embedding in LR White or Lowicryl K4M at 0 degrees C, or processing at progressively lower temperatures (PLT) and embedding in Lowicryl at -35/-50 degrees C. In a third strategy, specimens sensitive to very low fixative concentrations are cryo-immobilized, then resin embedded after substitution or freeze-drying (this latter method awaiting evaluation for inclusion in our strategical approach).

Acrylic Resins↗
PubMed1990

LR White embedding allows a multi-method approach to the analysis of brain tissue from patients with Alzheimer's disease.

Light (LM) and electron (EM) microscope comparisons of the cytochemistry and immunocytochemistry of neuritic plaques and neurofibrillary tangles, the main histopathological changes in the brains of Alzheimer's disease sufferers, have been almost impossible because of the disparity between the two technologies. By embedding unosmicated brain tissue in the acrylic resin LR White, direct comparisons can be made between techniques applied at the LM level with those at the EM level. After partial dehydration in 70% ethanol, the tissue is embedded by rapid infiltration and polymerization at 0 degrees C, which has been shown to maximally preserve tissue immunoreactivity. Semithin sections are then receptive to routine LM stains, silver stains e.g. Gomori's methenamine silver, and immunocytochemistry with immunoperoxidase or immunocolloidal gold. Serial thin sections are stable in the EM and can be immunolabelled and directly compared with their LM counterparts. Results from the use of a mouse monoclonal antibody against beta-amyloid and a rabbit polyclonal antibody against ubiquitin are presented. LR White resin includes no elements other than carbon, oxygen and nitrogen, of which it is composed, so that sections of it are valuable for sensitive X-ray energy dispersive microanalysis.

Acrylic Resins↗
PubMed1990

An immunocytochemical method for localization of estrogen receptors in rat tissues using a dinitrophenyl (DNP)-labeled rat monoclonal primary antibody.

We have developed an immunocytochemical method to demonstrate estrogen receptor in hormone-sensitive tissues of the rat using a dinitrophenyl (DNP) hapten-labeled rat antihuman estrogen receptor monoclonal antibody (MAb), H222. Mouse IgM anti-DNP was used secondarily, followed by a DNP/peroxidase conjugate, diaminobenzidine/hydrogen peroxide chromogen, and silver intensification. This method was applied to tissues from intact female rats and showed that estrogen receptor was localized in the nuclei of the stromal and glandular components of the uterine endometrium. Reduced receptor staining was observed in the luminal epithelium, with minimal myometrial staining. Anterior pituitary glands showed heterogeneous immunostaining and ovaries expressed the receptor predominantly in the interstitial cells; fallopian tubes demonstrated substantial epithelial staining. Uteri from chemically castrated rats showed reduced estrogen receptor immunostaining in both stromal and luminal cells, whereas staining was enhanced in the glandular elements. Classical estrogen-unresponsive tissues (heart, lung, and spleen) were unstained. Antibody controls involved pre-blocking antibody recognition sites on the receptor with unlabeled antibodies to estrogen receptor (H222, H226, and D547), as well as use of an inappropriate DNP-labeled antibody to metallothionein. These controls illustrated the specific nature of the DNP-H222 binding.

Adnexa Uteri↗
PubMed1989

The morphological and immunohistochemical analysis of renal biopsies by light and electron microscopy using a single processing method.

A methodology is described in which a number of well-established research techniques are brought together to enable the complete diagnostic analysis of a renal biopsy on a single piece of tissue. By embedding the biopsy in the acrylic resin LR White, unsupported sections of which are stable in the electron beam, light and electron microscopy and immunocytochemistry become feasible on sections from the same block. The biopsy is glutaraldehyde fixed but post-fixation in osmium tetroxide, which is often deleterious to antigen preservation, is omitted. Extraction in organic solvents and resin monomer is minimized by rapidly infiltrating the tissue from 70% ethanol and polymerizing the resin catalytically at 0 degrees C. Semithin sections can be stained with haematoxylin and eosin, Toluidine Blue or methenamine silver, giving results similar or superior to those obtained from paraffin sections. Thin sections show that the standard of morphological preservation is similar to that seen using epoxide sections even though the kidney is unosmicated. The tissue retains a high level of antigen reactivity, which, in the limited number of cases so far examined, has paralleled or exceeded that demonstrated by conventional immunofluorescence on frozen sections.

Biopsy↗
PubMed1989

Multiple hormone storage by cells of the human pituitary.

While immunostaining serial semi-thin sections of acrylic resin-embedded normal human pituitary using antisera to human pituitary hormones, it became clear that several cells were stained by more than one antiserum. The tissue had been surgically excised from a patient with a prolactinoma. The tumor, which was immunoreactive only with antiprolactin antiserum, was distinctly different from the pieces of tissue under study which had normal pituitary architecture and demonstrated immunoreactivity with antisera against all six of the common pituitary hormones. A major immunoelectron microscopic investigation, using immunocolloidal gold and immunoperoxidase methods, revealed cells in which follicle-stimulating hormone (FSH), luteinizing hormone (LH), and prolactin (PRL) were co-localized to the same electron-dense granules. Some similar cells also possessed electron-lucent granules immunoreactive only for anti-PRL antiserum. Adrenocorticotrophic hormone (ACTH) and PRL were also found in the same cell but were very largely localized to separate, morphologically different populations of electron-dense and -lucent storage granules. By employing double immunolabeling, a few granules in the ACTH/PRL cells were shown to be immunoreactive to both anti-ACTH and anti-PRL antisera. The possibility that the multipotential stem cells is discussed.

Adult↗
PubMed1987

Modern acrylics for post-embedding immunostaining techniques.

We describe two methods for rapid processing of biological tissues into LR White acrylic plastic. Both methods make use of LR White's compatibility with small amounts of water, enabling non-osmicated tissue to be only partially dehydrated before infiltration with the plastic, a procedure that improves the sensitivity of post-embedding immunocytochemistry. In addition, both methods are designed to reduce the time for which tissue is exposed to the damaging influence of the plastic monomer, which can cause extraction and sudden shrinkage. The tissue example used in the first method is immersion-fixed, surgically removed human pituitary which, by virtue of its thorough fixation, can be processed quickly at 50 degrees C using catalytic polymerization at room temperature. The concentration of the catalyst is critically set to prevent the temperature rising above 60 degrees C in the tissue blocks. Penetration of immunoperoxidase reagents into 330-nm LR White sections is demonstrated and possible modes of action are discussed. When "lightly" fixed tissue is processed as above, serious polymerization artifacts can result from autocatalysis. A second method, based on the first but employing slower polymerization at 0 degrees C, has therefore been developed. The high level of fine structure that can be retained using this method is illustrated by the demonstration of the trans-tubular Golgi in perfusion-fixed kidney of rat. Biotinylated lectin is localized to cells of the kidney proximal tubule with streptavidin-colloidal gold, to illustrate tissue reactivity. In a second example, the structure of the bacterial cell envelope is shown to be similar in appearance after partial dehydration and LR White embedding to that seen after progressive lowering of temperature, dehydration, and Lowicryl embedding.

Acrylic Resins↗
PubMed1986

Immunogold electron-microscopic localisation of calpain I in skeletal muscle of rats.

By using a double-affinity-purified first antibody and colloidal gold-conjugated second antibody, it is shown that calpain I (a cysteine proteinase activated by micromolar concentrations of Ca2+) has a predominant intracellular location in the I-band region of the extensor digitorum longus (EDL) muscle of the rat, but is not exclusively associated with the Z-line.

Animals↗
PubMed1986

Immunohistochemical localization of cell surface receptors using a novel method permitting simple, rapid and reliable LM/EM correlation.

A method is presented which allows correlative serial section analysis by light and electron microscopy of cell surface antigens in monolayer cultures. Sites of antigenicity are shown by deposition of diaminobenzidine after pre-embedding, immunoperoxidase immunocytochemistry. Osmication is replaced by the use of gold chloride which specifically enhances the electron density of diaminobenzidine. In addition gold chloride bound to diaminobenzidine survives embedding and provides the basis for a post-embedding photochemical amplification method. Immunostained cells are embedded in LR White by a rapid technique which preserves their structure and leaves them available for subsequent post-embedding immunocytochemistry. The method is illustrated by the demonstration of epidermal growth factor (EGF) receptors on the EGF receptor-rich human carcinoma cell line A431 using a well characterized monoclonal antibody raised against EGF receptor.

Antigens, Surface↗
PubMed1986

Immunohistochemical identification of prolactin and 24K protein in secretory endometrium.

Inadequate endometrial differentiation is a cause of infertility and recurrent pregnancy loss. Diagnostic histologic dating criteria may be supplemented by the immunohistochemical identification of protein markers in late secretory endometrium. Late secretory endometrium has been shown to contain prolactin. The authors report the immunohistochemical localization of prolactin and 24K protein in late secretory endometrium using monoclonal antisera and the dinitrophenyl hapten sandwich-staining technique. The appearance of these proteins in decidualized stromal cells of late secretory endometrium may provide a more specific indicator of endometrial development and differentiation, if their late appearance or absence can be correlated with pregnancy wastage.

Antibodies, Monoclonal↗
PubMed1984

Rat anterior pituitary cells maintained on artificial capillaries: responses of thyrotrophs and lactotrophs to depolarization, TRH and dopamine.

Rat anterior pituitary cells have been maintained over an 18-day period in a perfusion system designed around artificial capillaries. Using novel methodology the cells have been visualized by light microscopy and appear as aggregates, closely attached to and sometimes stretching around the capillaries. Their morphology is consistent with previous histology at the level of light microscopy. The techniques described are compatible with immunohistochemistry and electron microscopy. The functional integrity of thyrotrophs and lactotrophs maintained in the system has been examined by measuring the dynamics of TSH and PRL secretion in response to depolarization, TRH and dopamine (DA). TSH and PRL were significantly and reproducibly released by TRH over a 7-day period. On each day the release was dose-dependent with a threshold of at least 28 pg. Qualitatively the responses were rapid in onset (within minutes) for both hormones. Similar responses were measured in response to high K+ depolarization. Basal secretion of TSH and PRL was rapidly and significantly inhibited by DA in a dose-dependent manner (ED50 20 +/- 25 nM for TSH and 70 +/- 40 nM for PRL). Inhibition was dependent on the continued presence of DA and could be mimicked by bromocriptine and stereospecifically prevented by the active but not the inactive isomer of the DA receptor antagonist butaclamol. Simultaneous administration of 10(-6) M DA with 10(-8) M TRH prevented the release of TSH and PRL. The effect of DA was transient, subsequent TRH responses being qualitatively and quantitatively normal.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1984

Glandular duodenal carcinoid--a somatostatin rich tumour with neuroendocrine associations.

The clinical and pathological features of four cases of duodenal carcinoid tumour are presented. All four tumours showed a glandular pattern, and in three cases this was associated with psammoma bodies. In three tumours somatostatin was identified by immunocytochemistry in most tumour cells. In two cases the duodenal tumours were associated with von Recklinghausen's disease and phaeochromocytoma. The importance of these unusual features is discussed, and it is suggested that these glandular carcinoids are a specific subgroup of endocrine cell tumours which appear to have potentially important clinical and pathological associations.

Adrenal Gland Neoplasms↗