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PubMed1996

Detection of p53 gene abnormality by sequence analysis of archival paraffin tissue. A comparison with fresh-frozen specimens.

This parallel study was designed to compare the sensitivity and specificity of detection of point mutations in fresh-frozen and formalin-fixed, paraffin-embedded breast cancer tissue. Sequence analysis of exon 5 of p53 gene was performed on polymerase chain reaction-amplified DNA from 25 infiltrating ductal carcinomas of the breast. Four tumor showed mutations with identical base substitutions in their respective codons of both frozen and paraffin-embedded specimens. We conclude that subtle genetic alterations can be detected in archival paraffin tissue with an accuracy comparable to that of fresh-frozen histologic samples.

Breast Neoplasms↗
PubMed2005

On the art of anastomotic posterior urethroplasty: a 27-year experience.

PURPOSE: We determined the various operative details of anastomotic posterior urethroplasty that are essential for a successful result. MATERIALS AND METHODS: We reviewed the medical records of 155 patients who had undergone anastomotic repair of posterior urethral strictures or distraction defects between 1977 and 2003. Patient age ranged from 3 to 58 years (mean 21) and all except 1 had sustained a pelvic fracture urethral injury as the initial causative trauma. Repair was performed with a perineal procedure in 113 patients, elaborated perineal in 2 and perineo-abdominal in 40. Followup ranged from 1 to 22 years. RESULTS: The results were successful in 104 (90%) cases after perineal (including 2 elaborated perineal) and in 39 (98%) after perineo-abdominal repair. Successful results were sustained for up to 22 years after surgery. Urinary incontinence did not develop in any patients while 2 lost potency as a direct result of anastomotic surgery. CONCLUSIONS: Of the operative details 3 constitute the gold triad that assures a successful outcome, namely complete excision of scarred tissues, fixation of healthy mucosa of the 2 urethral ends and creation of a tension-free anastomosis. When the bulboprostatic urethral gap is 2.5 cm or less, restoration of urethral continuity may be accomplished with a perineal procedure after liberal mobilization of the bulbar urethra. For defects of 2.5 cm or greater the elaborated perineal or perineo-abdominal transpubic procedure should be used. In the presence of a competent bladder neck, anastomotic surgery does not result in urinary incontinence. Impotence is usually related to the original trauma and rarely (2%) to urethroplasty itself.

Adolescent↗
PubMed1980

Immuno-electron microscopical investigations with a new tracer: peroxidase-labeled protein A: application for detection of pemphigus and bullous pemphigoid antibodies.

Peroxidase-labeled Protein A, stable immunoenzyme tracer of high reactivity and comparatively low molecular weight, has been applied in immuno-electron microscopy for detection of bound IgG-type pemphigus and bullous pemphigoid antibodies. Comparing Protein A-peroxidase with peroxidase-labeled immunoglobulins, we obtained similar morphological results in corresponding incubation techniques, but lower nonspecific adsorption of Protein A-peroxidase complexes in tissues. The Protein A-peroxidase molecules showed good tissue penetration abilities. Our rapid one-step incubation procedure led to enhanced preservation of tissue fine structures, without the need of prior tissue fixation. It seems that Protein A-peroxidase is able to replace peroxidase-labeled anti-IgG for immuno-electron microscopical purposes.

Humans↗
PubMed1994

Comparison of two methods of storing breast fine needle aspirates (FNAs) using oestrogen receptor immunocytochemical assay as a method of evaluating the storage methods.

Two methods of storing fine needle aspirates were compared in 14 patients with breast cancer. The methods of storage were: (1) as a Cytospin slide prepared immediately from the aspirated material and stored at -80 degrees C; (2) as a suspension of cells in tissue culture medium, stored at -80 degrees C. The effect of storage on the cells was assessed by means of an oestrogen receptor immunocytochemical assay (ER-ICA). An ER positivity of 100% was obtained by ER-ICA staining of cells after storage method 1, whilst all of the specimens stored by method 2 were ER-negative. The data demonstrate that cells stored in tissue culture medium at -80 degrees C are not suitable for ER measurement. The storage method of choice for specimens intended for ERICA is as a Cytospin slide. The ER status of cells deposited on Cytospin slides prepared immediately and stored at -80 degrees C for 2 years could be demonstrated despite the delay in processing the specimen.

Biopsy, Needle↗
PubMed1995

Human endothelial cells bioactivate organic nitrates to nitric oxide: implications for the reinforcement of endothelial defence mechanisms.

Although in therapeutic use for more than a century, the mode of cellular action of organic nitrates remains incompletely understood. Despite ample experimental evidence from animal studies to show that nitrates are metabolized to NO in the vascular smooth muscle, direct demonstration of such an activity in human vascular cells is still lacking. Moreover, the role of the endothelium in modulating the pharmacodynamic action of nitrates is far from clear. We therefore aimed to investigate whether or not human endothelial cells are capable of bioactivating these drugs to NO and whether the amounts generated are sufficient to elicit any biological effects. Using cultured human umbilical vein endothelial cells (HUVECs) as an established model system a combination of three different methods was used to address this issue: (1) quantification of NO formation upon endothelial nitrate metabolism using the oxyhaemoglobin technique; (2) evaluation of the second messenger response using radioimmunoassay for cGMP; and (3) assessment of mechanism and extent of potentiation of the anti-aggregatory effect of nitrates in the presence of endothelial cells as a relevant bioassay. We now show that superfusion of cultured human endothelial cells on microcarrier beads with either glyceryl trinitrate (GTN) or isosorbide dinitrate (ISDN; both at 0.1-100 mumol L-1) results in a concentration-dependent formation of NO. NO generation from isosorbide 5-mononitrate (IS-5-N) was below the detection limit. The amounts of NO produced (maximally 2.97 +/- 0.98 pmoles NO min-1 x mg protein with 100 mumol L-1 GTN; n = 8) were similar to those elicited upon challenge of the cells with 100 nM bradykinin. NO formation from either organic nitrate was accompanied, in a concentration-dependent and methylene blue-inhibitable manner, by stimulation of endothelial soluble guanylyl cyclase with consequent increases in the intracellular level of cGMP (maximally 32-fold over basal levels with ISDN), a significant portion of which was released into the extracellular space. Upon continuous 30 min superfusion or repeated application of high concentrations of GTN (100 mumol L-1) nitrate bioactivation to NO was subject to partial tachyphylaxis. Co-incubation of washed human platelets with HUVECs potentiated the anti-aggregatory action of nitrates in a cell number dependent and oxyhaemoglobin-sensitive manner and this effect, too, was accompanied by increases in intraplatelet cGMP levels. The potentiating effect was largely inhibited after blockade of sulfhydryl groups by pre-incubation of HUVECs with N-ethylmaleimide and completely abrogated after pretreatment of cells with the tissue fixative glutaraldehyde. These results demonstrate that human endothelial cells are capable of bioactivating organic nitrates to NO by an enzymatic, apparently thiol-sensitive pathway, in quantities sufficient to influence endothelial and platelet function. Besides the well known vasorelaxant action of organic nitrates, which is mainly due to their metabolism in the smooth muscle compartment, these drugs may therefore be endowed with a hitherto underestimated potential to directly influence endothelial functions via the NO/cGMP pathway. Through specific bioactivation in the endothelium itself organic nitrates can thus mimic and reinforce protective functions normally served by a functional endothelium such as the modulation of blood cell/vessel wall interactions and inhibition of cell proliferation.

Biotransformation↗
PubMed1994

Detection of neutral endopeptidase (NEP, enkephalinase, E.C.3.4.24.11) in relation to dopaminergic and gonadoliberinergic nerve endings in the median eminence of the male rat: a double labeling ultrastructural study.

The existence of neutral endopeptidase (Enkephalinase, NEP, E.C.3.4.24.11) in membranes of nerve endings in the rat median eminence suggests that some neuropeptides have paracrine and/or autocrine actions in this region. In vitro, neutral endopeptidase is capable of hydrolysing a variety of regulatory peptides but in vivo, many works indicate that in the central nervous system this enzyme is highly implicated in the biological inactivation of enkephalins and tachykinins. In addition there is evidence that NEP is also involved in the inactivation of neurotensin in vivo. The modulation of the release of gonadotrophin releasing hormone (GnRH) is one of the documented actions of enkephalins within the median eminence. However, it is at present unclear whether enkephalins act on dopamine endings, on GnRH endings or on both. As the technical parameters and particularly the tissue fixation used to detect neutral endopeptidase are compatible with immunocytochemical detection of GnRH and tyrosine-hydroxylase (the rate limiting enzyme in the synthesis of catecholamines), two double immunolabelings were realised at the ultrastructural level to determine if GnRH and dopamine nerve endings have the enzyme inserted within their plasma membrane. Our study shows the presence of neutral endopeptidase on tyrosine-hydroxylase-immunoreactive nerve endings while presence of the enzyme on GnRH-immunoreactive nerve endings is not demonstrated. Consequently, our results provide morphological arguments for possibilities of paracrine and/or autocrine actions by neuropeptides inactivated by neutral endopeptidase on tuberoinfundibular dopaminergic nerve endings. Conversely, action of the same peptides on GnRH boutons seems more unlikely.

Animals↗
PubMed1978

Subcellular metabolite concentrations. Dependence of mitochondrial and cytosolic ATP systems on the metabolic state of perfused rat liver.

Mitochondrial and cytosolic contents of adenine nucleotides and phosphate were measured in perfused rat livers employing a technique of fractionation of freeze-fixated tissue in non-aqueous solvents. From the subcellular contents the mitochondrial and cytosolic concentrations of ATP, ADP, AMP and phosphate and the phosphorylation potentials of the subcellular ATP systems were calculated. An attempt was made to elucidate the relationship between mitochondrial and cytosolic adenine nucleotide systems and the dependency on the metabolic state of the liver. The following results were obtained: 1. Under all metabolic conditions studied the mitochondrial ATP/ADP ratios were considerably lower than the cytosolic ratios (mitochondria: 0.1-0.7; cytosol: 2-11). 2. The ATP/ADP ratios calculated from overall tissue contents reflect mainly the cytosolic ratios. 3. An inverse relationship was found between mitochondrial and cytosolic ATP/ADP ratios, i.e. when the mitochondrial ratios tended to increase, the cytosolic ratios decreased and vice versa. 4. The phosphorylation potentials calculated from the subcellular concentrations were higher in the cytosol than in the mitochondria. The potential difference varied between 11 and 3 kj/mol in livers from fed and starved rats, respectively. 5. In the presence of mitochondrial inhibitors, i.e. amytal, dinitrophenol and carboxyatractyloside, the potential difference between the subcellular ATP systems decreased predominantly due to an increase in the mitochondrial ATP/ADP ratios. 6. A correlation between mitochondrial ATP/ADP ratios and the respiratory rates was not observed, but the subcellular ratios appeared to correlate with the rate of glycolysis. When the rate of lactate + pyruvate production was increased, the cytosolic ATP/ADP ratios were increased, too, whereas the mitochondrial ratios tended to decrease. 7. The adenine nucleotides in the cytosol appear to be in near equilibrium catalysed by the adenylate kinase. In the mitochondria, the AMP concentration is much lower than to be expected under equilibrium conditions. These results were discussed with respect to rate control of processes involved in ATP generation, i.e. oxidative phosphorylation, adenine nucleotide translocation and glycolysis.

Adenosine Diphosphate↗
PubMed2004

Proliferative glomerulonephritis with monoclonal IgG deposits: a distinct entity mimicking immune-complex glomerulonephritis.

BACKGROUND: Renal disease related to the deposition of monoclonal immunoglobulins containing both heavy and light chains can occur in type 1 cryoglobulinemia, Randall type light and heavy chain deposition disease (LHCDD), and immunotactoid glomerulonephritis. We report a novel phenotype of glomerular injury that does not conform to any of the previously described patterns of glomerular involvement by monoclonal gammopathy. METHODS: Ten cases of unclassifiable proliferative glomerulonephritis manifesting glomerular monoclonal immunoglobulin G (IgG) deposits were identified retrospectively from the archives of the Renal Pathology Laboratory of Columbia University over the past 3 years (biopsy incidence 0.21%). RESULTS: The monoclonal immunoglobulins formed granular electron dense deposits in mesangial, subendothelial, and subepithelial sites, mimicking ordinary immune complex-mediated glomerulonephritis and producing a diffuse endocapillary proliferative or membranoproliferative glomerulonephritis. However, by immunofluorescence, the deposits were monoclonal, staining for a single light chain isotype and a single gamma subclass (including two IgG1kappa, one IgG1lambda, one IgG2lambda, four IgG3kappa, and one IgG3lambda). All cases stained for the three constant domains of the gamma heavy chain (CH1, CH2, and CH3), suggesting deposition of a nondeleted immunoglobulin molecule. Tissue fixation of complement was observed in 90% of cases, and 40% of patients had hypocomplementemia. Clinical presentations included renal insufficiency in 80% (mean serum creatinine 2.8 mg/dL, range 0.9 to 8.0), proteinuria in 100% (mean urine protein 5.8 g/day; range 1.9 to 13.0), nephrotic syndrome in 44%, and microhematuria in 60%. A monoclonal serum protein with the same heavy and light chain isotype as that of the glomerular deposits was identified in 50% of cases (including three IgGkappa and two IgGlambda); however, no patient had clinical or laboratory features of type 1 cryoglobulinemia. No patient had overt myeloma or lymphoma at presentation or over the course of follow-up (mean 12 months). CONCLUSION: Glomerular deposition of monoclonal IgG can produce a proliferative glomerulonephritis that mimics immune-complex glomerulonephritis by light and electron microscopy. Proper recognition of this entity requires confirmation of monoclonality by staining for the gamma heavy chain subclasses.

Adult↗
PubMed1986

Benign monoclonal IgAK gammopathy associated with polyneuropathy and dysautonomia.

The first case of benign IgAK monoclonal gammopathy associated with peripheral neuropathy is described. Dysautonomia is an unusual, yet prominent, manifestation of neuropathy in this patient. Electrodiagnostic testing and nerve biopsy were compatible with demyelination and axonal loss. Myelin sheath, perineural, and endoneural interstitial tissue fixation of anti-IgA and anti-kappa light chains was demonstrated by direct immunofluorescence microscopy. Absorption studies utilizing human peripheral nerve myelin resulted in complete removal of the paraprotein band. Analytic procedures with myelin-associated glycoprotein and gangliosides, however, were negative. Based on these findings, an alternative etiology for this neuropathy is hypothesized.

Aged↗
PubMed1997

In situ hybridization study on tissue inhibitors of metalloproteinases (TIMPs) mRNA-expressing cells in human inflamed gingival tissue.

This study presents the exact cell types and localization of tissue inhibitors of metalloproteinases (TIMPs) production sites in periodontal diseased gingiva by means of in situ hybridization. Gingival tissue specimens were fixed, embedded and hybridized in situ with specific digoxigenin-labeled cRNA probes (386 and 496 bp). TIMP-1 and -2 mRNAs were expressed on macrophages, mononuclear cells, capillary endothelial cells and some fibroblasts throughout the gingival tissue. In periodontitis, TIMP-1 and -2 mRNA-expressing cells showed significantly different localization. TIMP-1 mRNA was broadly observed in the gingival connective tissue while TIMP-2 mRNA was predominantly expressed in the connective tissue adjacent to the pocket epithelium (p < 0.01). Fewer TIMPs mRNA were observed in minimal gingivitis than in periodontitis, especially in the middle zone of gingival tissue. Thus, TIMP-1 and TIMP-2 mRNA was detected differentially and site-specifically in periodontal diseased gingival tissue.

Aged↗
PubMed1984

Studies on hydragogue drugs: light and electron microscopic examination of the isolated rat colonic mucosa exposed to deoxycholic acid and synthetic surfactants.

Sacs of the stripped and everted, isolated descending rat colon were incubated for 2 hours in presence of the following surfactants at the mucosal side: Dodecylsulphate (DDS), dioctylsulphosuccinate (DOSS), cetrimonium bromide (CTMAB), Triton X100 and deoxycholic acid (DOC). After tissue fixation, the sacs were processed for light microscopy (LM) and for scanning (SEM) and transmission (TEM) electron microscopy. All three methods revealed that DOSS (1.3 X 10(-4) and 2.6 X 10(-4) mol/l, CTMAB (5 X 10(-5) and 1 X 10(-4) ) and Triton (2 X 10(-5), 5 X 10(-5) and 1 X 10(-4) ) caused only minor or moderate changes compared to parallel controls, as did also DDS at 1 X 10(-5) and 2 X 10(-5) mol/l. DDS at 2 X 10(-4) and 4 X 10(-4) mol/l and DOC at 1.5 X 10(-4) and 3 X 10(-4) mol/l caused more prominent changes. LM showed swollen, vacuolated cells with pycnotic nuclei; many of these cells seemed to be extruded. According to SEM, cells thus affected were most abundantly localized to the normal extrusion zone at the borders of the crypt-surface epithelial cell units. DOC tended to cause a more generalized affection within the units than DDS. In spite of these deleterious effects, gaps corresponding to missing epithelial cells were not observed. TEM indicated the mechanism responsible for restoration of epithelial continuity in spite of extensive cell loss: The remaining epithelial cells seemed to flatten out and re-establish cell-to-cell contact by pseudopod formation along the basement lamina. This repair mechanism seemed to operate at a rapid rate; however, incomplete closure of cellular gaps i.e. small denuded parts of the basement lamina were occasionally observed. The results of this study are discussed in relation to a functional study under identical experimental conditions (Gastroenterol. Clin. Biol. 1981, 5, 124), in which these surfactants caused a significant alteration of normal colonic transport function.

Animals↗
PubMed1998

Equine laminitis basement membrane pathology: loss of type IV collagen, type VII collagen and laminin immunostaining.

Disintegration of the basement membrane (BM) of the equine hoof lamellae and failure of the BM to remain attached to the basal cells of the secondary epidermal lamellae (SEL) is one of the earliest pathological events to occur in acute laminitis. Changes in the lamellar basement membrane were investigated by immunolabelling the key structural components of the BM, type IV collagen, type VII collagen and laminin in the lamellar BM of horses 48 h after the induction of laminitis. Lamellar tissues were harvested from 2 normal horses and 2 horses with acute laminitis. Immunostaining with antibody raised against human epitopes for type IV collagen, type VII collagen and laminin successfully stained the basement membranes of horse hoof lamellar tissues. Vascular tissue did not immunostain with type VII collagen antibody. Normal BM stained as a fine dark brown line and the lamellar BM was adhered to the basal cells of the SELs with no evidence of lamellar separation. At least 2 changes to the lamellar BM occurred in acute laminitis: loss of attachment of lamellar epidermal basal cells to their underlying BM and disintegration of the lamellar BM. In some sections from feet affected by acute laminitis, there was widespread separation of the SELs from their BM without loss of BM immunostaining and in others there was extensive loss of BM immunostaining. In lesions characterised by lamellar separation, the epidermal basal cells at the tips of the primary epidermal lamellae appeared to have slipped away from their BM and were an amorphous clump of epidermal cells devoid of immunostained BM. The BM from which they had separated remained in its original position in the dermis and was clearly outlined by all 3 antibodies. In other areas, however, virtually all the BM immunoreactivity at the PEL tips was absent. Only the occasional distorted SEL tip and fragments of BM retained sufficient immunostaining to allow anatomical identification. Numerous polymorphonuclear leucocytes (PMNs) invariably surrounded the tips of lamellae showing large scale loss of immunoreactivity and many PMNs had penetrated the lamellar BM and were within the epidermal compartments. PMNs were less frequent in the midlamellar region. Immunostaining of the BM of many SELs was absent in the midlamellar region. In some lamellae loss of BM immunostaining had occurred only at the bases of the SELs and fragments of immunostained BM were present in the zones of lysed BM suggesting that BM lysis was incomplete at the time of tissue fixation. In other lamellae, lysis of the BM was complete; there was no immunostained BM between SELs and the bulk of the epidermal cells of each PEL were an amorphous column of cells on either side of the central keratinised axis of the PEL. The lamellar BM which remained appeared as immunostained strands of unattached BM along the edges of the PDLs. Activation of BM degrading metalloproteinases (MMPs) occurs in acute laminitis and it seems likely that uncontrolled MMP activity is responsible for the loss and disorganisation of lamellar BM demonstrated in this study.

Acute Disease↗
PubMed1990

Kidney: a target organ for calcitonin gene-related peptide.

In the present study we have evaluated the effect of calcitonin gene-related peptide (CGRP) on cyclic AMP levels in intact mouse kidneys. We have used an in vivo bioassay based on microwave irradiation as a means of rapid tissue fixation, followed by the determination of cyclic AMP concentration using a protein binding assay. CGRP was found to cause a dose-dependent elevation of renal cyclic AMP levels, and despite being slightly less potent than calcitonin (CT), the action of CGRP was significantly more prolonged. The extended time course of action of CGRP is in sharp contrast to its known effect on bone cyclic AMP levels and could be of physiological relevance. CGRP may act on a receptor different from that of CT to produce distinct functional effects.

Animals↗
PubMedDate not supplied

Seeing the invisible: the challenge of imaging vitreous.

Imaging the vitreous is an attempt to view what is by design invisible. The inability to adequately image vitreous hinders a more complete understanding of its normal structure and function and how these change in aging and disease. The combined use of more than one technique could provide better imaging for investigational and clinical purposes. Past and present imaging methodologies are summarized and research and clinical techniques that are currently in development for future applications, are discussed. Dark-field slit microscopy has been used to characterize vitreous anatomy, both within the vitreous body as well as at the vitreo-retinal interface. In addition to this methodology, slit-lamp biomicroscopy; direct, indirect, and scanning laser ophthalmoscopies; ultrasonography; optical coherence tomography; magnetic resonance and Raman spectroscopies; and dynamic light-scattering methodologies for noninvasive evaluation are presented. Dark-field slit microscopy enables in vitro imaging without dehydration or tissue fixatives. Optical coherence tomography enables better in vivo visualization of the vitreo-retinal interface than scanning laser ophthalmoscopy and ultrasonography, but does not image the vitreous body. Dynamic light scattering can determine the average sizes of vitreous macromolecules within the vitreous body as well as possibly image the posterior vitreous cortex once detached, while Raman spectroscopy can detect altered vitreous molecules, such as glycated collagen and other proteins in diabetic vitreopathy.

Eye Diseases↗
PubMed1992

Effects of prolonged processing in standard and isotonic trichloroacetic acid (TCA) on cellular preservation in bone marrow trephines.

The effects of a combined fixing and decalcifying solution, trichloroacetic acid, on prepared bone marrow specimens were evaluated to try to improve the speed of trephine biopsy processing in our laboratory. The morphological preservation obtained immediately after decalcification was complete was assessed, and also after prolonged immersion in each of two solutions. Trichloracetic acid was made up in the standard manner using distilled water and in an isotonic form by the addition of sodium chloride. Prolonged immersion was performed to assess any possible adverse effects of this acidic solution. By the time decalcification was complete, each solution gave morphological preservation sufficient for diagnostic purposes. However, after prolonged immersion, only the isotonic solution performed adequately.

Biopsy↗
PubMed2000

Limitations of clonality analysis of B cell proliferations using CDR3 polymerase chain reaction.

BACKGROUND/AIMS: Detection of clonal immunoglobulin heavy chain (IgH) rearrangements by the polymerase chain reaction (PCR) is an attractive alternative to Southern blotting in lymphoma diagnostics. However, the advantages and limitations of PCR in clonality analysis are still not fully appreciated. In this study, clonality was analysed by means of PCR, focusing in particular on the sample size requirements when studying extremely small samples of polyclonal and monoclonal lesions. MATERIALS/METHODS: High resolution complementarity determining region 3 (CDR3) PCR was used to investigate the minimum number of cells and the amount of tissue required for the detection of a polyclonal population, both for fresh cells and formalin fixed, paraffin wax embedded tissue. Subsequently, frozen and paraffin wax embedded samples of 76 B cell lymphoproliferative disorders, 43 of which were tested by means of Southern blotting, were analysed to establish the sensitivity of this assay. These specimens included 12 chronic lymphocytic leukaemias (CLLs), nine mantle cell lymphomas (MCLs), 10 follicular lymphomas (FLs), and 45 mucosa associated lymphoid tissue (MALT) lymphomas. The specificity was tested on reactive lymph nodes (n = 19), tonsils (n = 4), peripheral blood lymphocyte fractions (n = 4), and biopsies with gastritis (n = 21). RESULTS: In reactive tissue, 20 ng of high molecular weight DNA derived from 6.5-9 x 10(3) B cells was sufficient to obtain a polyclonal PCR result. With smaller amounts "pseudoclonality" could be induced. When using paraffin wax blocks, undiluted DNA isolated from tonsillar tissue of at least 1 mm2 was necessary to obtain a polyclonal pattern. The sensitivity required to detect clonality in paraffin wax embedded and frozen tissue by PCR for FL (40% and 60%, respectively) was lower than that for MALT lymphomas (60% and 86%, respectively), CLL (78% and 89%, respectively), and MCL (88% and 100%, respectively). PCR specificity was 96% and 100% for frozen and paraffin wax embedded tissue, respectively. CONCLUSION: The minimum amount of template for CDR3 PCR is approximately 20 ng of high molecular weight DNA or 1 mm3 of B cell rich paraffin wax embedded normal tonsillar tissue, but care has to be taken to avoid pseudoclonality when low numbers of B cells are present. Duplicate or triplicate tests should be performed to avoid misinterpretation. The specificity of the PCR assay is almost 100%, whereas sensitivity depends on a combination of factors, such as lymphoma type and tissue fixation. Because frozen samples yield better results, obtaining fresh material for the PCR assay is recommended, especially when analysing FL and MALT lymphomas.

Blotting, Southern↗
PubMed1987

Synthetic ossicular replacements: normal and abnormal CT appearance.

Twenty-two patients with synthetic ossicular replacements were studied with computed tomography (CT). Twelve patients had total ossicular replacement prostheses (TORPs), and ten patients had partial ossicular replacement prostheses (PORPs). Good results were achieved in 12 patients (eight with TORPs and four with PORPs). Four of these prostheses were clearly visible on CT scans, six were discernible, and two were not identified. In the ten patients with unsatisfactory surgical results, CT was performed before surgical revision. It disclosed subluxation of the prosthesis in four patients and fibrous tissue fixation in four. In two patients the CT scan appeared normal; surgery disclosed lateralization of the graft with a nonfunctioning interface.

Ear Ossicles↗
PubMed1988

Cytochalasin B inhibition of toad bladder apical membrane responses to ADH.

The possible role of actin microfilaments in antidiuretic hormone (ADH)-induced increases in apical membrane water permeability was investigated in studies that evaluate inhibition by cytochalasin B of both permeability and membrane structural responses in the toad urinary bladder. Experiments were carried out in the absence of a transepithelial osmotic gradient to eliminate possible flow-induced distortions of the response. Measurements of osmotic water permeability after a brief tissue fixation with glutaraldehyde show that cytochalasin B reduces the permeability response to ADH by approximately one-third. Freeze-fracture electron microscopy indicates that the intramembrane particle aggregates, previously found to correlate closely with ADH-induced permeability, are reduced by about the same extent (28%) under these conditions. However, the frequency of apical membrane fusion events was not affected by cytochalasin B treatment. These results suggest that cytochalasin B treatment in the absence of an osmotic gradient alters the ADH-induced permeability through an effect on apical membrane aggregate frequency.

Animals↗