Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Tissue Fixation”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 343 records · Page 19Linked to original sources
PubMedDate not supplied

Improved hepatic and pancreatic localisation of the equine alpha-1-proteinase inhibitor family of serpins using an antigen enhancement technique and a monoclonal antibody.

Equine alpha-1-proteinase inhibitor (API) consists of three, occasionally four, serum glycoproteins. This study investigated the immunohistochemical localisation of equine API in paraformaldehyde fixed, paraffin embedded equine tissue samples of liver, lung, stomach, pancreas, jejunum and colon in five horses using affinity purified sheep polyclonal and protein A purified mouse monoclonal antibodies, whose specificities were verified by Western blotting. Exposing tissue sections to boiling citrate buffer greatly enhanced antigen recovery and improved immunostaining with both antibodies, resulting in discovery of novel tissue distribution patterns for the horse. In the horses studied, all hepatocytes showed some degree of cytoplasmic staining, many having perinuclear intense granular inclusions. This finding is contrary to findings in human studies where hepatocytes of Pi MM phenotype have proven difficult to stain for human API, despite evidence at the molecular level suggesting hepatocytes as the major source of serum API. This discrepancy may be due to the use of different tissue fixation and antigen recovery techniques. In all other tissues examined, the distribution of equine API was similar to human studies.

Animals↗
PubMed1987

Histophotometrical measurements concerning the distribution of myofibrillar ATPase activity within the tissue block after aldehyde fixation.

The inhibition of myofibrillar ATPase activity by aldehyde fixation in tissue pieces of myocardium of the rat was measured by scanning histophotometry. The relative amount of the final reaction product of the histochemical method for ATPase (Padykula and Herman 1955) was taken as a measure for the enzyme activity. Along scanning lines in slides from the surface to the centre of tissue block, a higher activity was found in the centre in contrast to the marginal zone. Compared with the centre of the block (approximately equal to 100%), in the marginal region a loss of about 40% (in paraformaldehyde PFA) and of 75% (in glutaraldehyde GA) was found, the latter with a sharp decline between the marginal zone and the central part. For quantitative enzyme histochemistry by histophotometry unfixed material is recommended.

Adenosine Triphosphatases↗
PubMed2002

Evans Blue Dye as an in vivo marker of myofibre damage: optimising parameters for detecting initial myofibre membrane permeability.

Evans Blue Dye (EBD) is widely used to study cellular membrane permeability and has recently been utilised in mdx mice to identify permeable skeletal myofibres that have become damaged as a result of muscular dystrophy. EBD has the potential to be a useful vital stain of myofibre permeability in other models of skeletal muscle injury and membrane-associated fragility. The parameters for its use for such purposes were optimised in the present study, of particular interest is the use of EBD to identify the onset of muscle damage. This study compared intravenous vs. intraperitoneal injection; tissue fixation; volume of EBD; time of availability in tissue; and persistence after injection in mdx mice (with endogenous muscle damage) and control mice. Satisfactory labelling of permeable myofibres was seen in frozen sections viewed with fluorescence microscopy when intraperitoneal injection of a 1% EBD solution injected at 1% volume relative to body mass was administered between 16 and 24 h prior to tissue sampling. EBD labelling was then assessed in three mouse models of experimental injury and repair-cut injury, whole muscle grafts, and exercise-induced muscle damage. These experiments demonstrated that (i) following a cut injury across myofibres, EBD penetrated up to 150 microm from the injury site over a 20-h period; (ii) EBD was present throughout myofibres of avascular whole muscle graft by one day after transplantation; and (iii) damaged myofibres were detected within 20 min after controlled lengthening-contraction exercise. This simple and inexpensive technique has sensitivity for the detection of increased myofibre permeability and/or sublethal damage that has advantages over other traditional histological techniques at the light microscopy level.

Animals↗
PubMed2001

Lack of association of prostate carcinoma nuclear grading with prostate specific antigen recurrence after radical prostatectomy.

PURPOSE: Grading prostate cancer using the Gleason system relies only on architectural tumor growth, in contrast to other systems, such as the WHO system, which grade prostate carcinoma based on nuclear features as well as architectural patterns. The prognostic significance of nuclear grading remains controversial since most studies were performed before prostate specific antigen (PSA) screening became widely available. We evaluated the significance of nuclear grade for predicting PSA recurrence in a contemporary cohort of patients treated with radical prostatectomy for clinically localized prostate carcinoma. MATERIALS AND METHODS: Nuclear grades 1 to 3 were determined in 141 consecutive radical prostatectomies in 1995. Predominant and worst nuclear grade was determined by a consensus of 3 pathologists. Statistical analysis compared nuclear grade with Gleason score using the chi-square test. The Cox proportional hazards analysis was performed to calculate the ability of nuclear grade, Gleason score and other variables to predict PSA recurrence. RESULTS: We identified a significant association of Gleason score with worst nuclear grade (p = 0.007). All 6 cases with a Gleason score of 8 or greater had a worst nuclear grade of 3, in contrast to 36 of 60 (60%) with a score 6 or less, in which the worst nuclear grade was 3. Of the 141 patients 31 (21.9%) had PSA recurrence at a median followup of 3.7 years. The univariate Cox model revealed significant associations of PSA recurrence with Gleason score 8 or greater (hazards ratio 5.5, p = 0.005), extraprostatic extension (hazards ratio 3.4, p = 0.001), positive surgical margin (hazards ratio 2.6, p = 0.009), seminal vesicle involvement (hazards ratio 7.3, p <0.001), preoperative serum PSA (hazards ratio 1.03, p = 0.007), tumor stage (hazards ratio 3.6, p = 0.001) and maximal tumor dimension (hazards ratio 2.4, p <0.001). However, overall and worst nuclear grade did not predict PSA recurrence (p = 0.89 and 0.13, respectively). Nuclear grade did not fit any multivariate model tested, which otherwise included Gleason score, log(PSA), surgical margin status, extraprostatic extension, seminal vesicle status, tumor size and pathological stage. By varying sample fixation time we also showed that benign prostate tissue in the same section as prostate carcinoma had grade 2 or 3 nuclear changes, that is moderate to marked anaplasia. CONCLUSIONS: High nuclear grade is associated with high Gleason score. However, prostate carcinoma with a Gleason score of 6 or less shows extreme variability. Nuclear grade determined by light microscopy failed to predict PSA recurrence in a contemporary series of men with clinically localized prostate cancer treated with radical prostatectomy. Nuclear morphology is subject to tissue fixation and processing artifact. Any nuclear morphometric study must consider this artifact.

Adult↗
PubMed2005

The influence of physical and chemical agents on photooxidation of porcine pericardial collagen.

Photooxidation is a method of tissue fixation resulting in protein crosslinking due to illumination in the presence of a dye. The aim of the study was to evaluate the impact of dyes, photooxidation time and the type of applied light on the porcine pericardial collagen crosslinking. The collagen modifications were evaluated on the basis of pericardial sensitivity to pepsin digestion. The hydrolysate components were evaluated qualitatively and quantitatively. All hydrolysates contained collagen alpha chains, their aggregates and degradation products. Methylene blue and methylene green-mediated 4 h photooxidation in the presence of visible light caused similar decrease in pericardium sensitivity to pepsin. However, both fixation types generated remarkable amounts of alpha chain degradation products. The prolongation of photooxidation time to 8 h did not increase the pericardial sample resistance to pepsin. Moreover, these sample hydrolysates revealed an elevated alpha chain content. Violet light mediated photooxidation did not alter pericardial sensitivity to pepsin when compared with fixation under visible light. Nevertheless, violet light fixed tissues displayed a decrease in collagen degradation products. The application of violet light in photooxidation of porcine pericardium will probably allow to obtain enzyme resistant bioprostheses with better mechanical properties compared with those obtained after visible light mediated process.

Animals↗
PubMed1985

Demonstration of myoglobin and CK-M in myocardium. Comparison of five fixation methods and three immunohistochemical techniques.

The results of immunohistochemical staining vary depending on the tissue, fixative, antigen-antibody system, and immunohistochemical staining methods used. The purpose of this study was to evaluate the effect of different methods of fixation, different antigen-antibody systems, and different immunohistochemical methods on immunohistochemical staining of myocardium. Samples of normal fresh canine myocardium from six dogs were fresh frozen and fixed in 10% neutral buffered formalin, Bouin's, Bayley's and Carnoy's fixatives. Immunohistochemical staining for myoglobin and creatine kinase M was performed using the ABC (avidin-biotin complex) and indirect peroxidase-antiperoxidase (PAP) techniques. Tissues fixed in formalin showed the most intense specific staining for both antigens with the least background and nonspecific staining. All other fixation methods and frozen section techniques gave a more variable degree of specific positive staining and substantial background staining and/or nonspecific staining. ABC and PAP techniques gave similar results with both antigen-antibody systems and with each fixation method. Thus, no differences in specificity or sensitivity were observed between ABC and PAP techniques. Differences in staining intensity and pattern were related primarily to differences in fixation methods.

Animals↗
PubMed1985

Vesicular profiles in frog perineural cells preserved by rapid-freezing and freeze-substitution.

Vesicular profiles in perineurial cells of frog peripheral nerves were examined in tissues preserved by rapid-freezing or by conventional chemical fixation. Tissues were processed immediately after removal from the animal or after remaining in Ringers for several hours. Vesicular profiles were present in perineurial cells in all experimental groups, demonstrating that they are not an artifact of chemical fixation. However, variations in their morphology correlated with the different preservation techniques.

Animals↗
PubMedDate not supplied

Differences in acetylcholine but not choline in brain tissue fixed by freeze fixation or microwave heating.

Among the many rapidly metabolized compounds in the brain, acetylcholine is one of the most challenging to sample effectively due to its rapid synthesis, degradation and sequestration. To ascertain problems that invalidate sampling procedures two methods of tissue fixation, microwave heat inactivation and freeze fixation, were used for obtaining mice and rat brain samples, respectively. The data show that acetylcholine levels obtained by microwave fixation were much higher than those obtained by freeze fixation. Choline levels were not affected by the fixation method used. Microwave fixation results in more accurate assessment of acetylcholine levels than the freeze fixation method, even though the tissue fixation time was less than 1 s in both methods, because tissue integrity is maintained in the microwave fixation, but not during freeze fixation.

Acetylcholine↗
PubMed1984

A comparison of the avidin-biotin-peroxidase complex (ABC) and peroxidase-anti-peroxidase (PAP) immunocytochemical techniques for demonstrating Sendai virus infection in fixed tissue specimens.

Mice were infected with Sendai virus and killed 8 days later. Lungs were removed and perfused with ethanol, 10% neutral formalin, Bouin's, B-5, or Zenker's fixatives. Tissues were dehydrated, embedded in paraffin, sectioned and stained for the presence of Sendai virus using the avidin-biotin-peroxidase-complex (ABC) and peroxidase antiperoxidase (PAP) immunocytochemical techniques. Results of these techniques were compared. The ABC technique was more sensitive than the PAP. Sendai antigen was demonstrated by the ABC technique in lung tissue fixed with any fixative, whereas antigen could be demonstrated with consistency only in ethanol-fixed lung by the PAP technique. Trypsin treatment of lung prior to immunoperoxidase treatment failed to enhance staining with either technique and actually caused a decrease in staining in ethanol, B-5 and Zenker's-fixed specimens.

Animals↗
PubMed1989

Astrocytes and microglia in human brain share an epitope recognized by a B-lymphocyte-specific monoclonal antibody (LN-1).

A B-lymphocyte-specific mouse monoclonal antibody, LN-1, recognizes two morphologic classes of glial cells in human brain. The nature and duration of tissue fixation and processing are critical in the detection of the two cell types. In tissue that is lightly fixed, LN-1 recognizes astrocytes. The astrocytic nature of the LN-1 reactive glial cell was confirmed by cytologic features, tissue distribution, immunoelectron microscopy, double labeling immunofluorescent microscopy, and staining of serial sections with antibodies to glial fibrillary acidic protein. In tissue that is fixed for longer periods or in Bouin's fixative, two glial cell types are recognized: astrocytes and microglia. The identity of the latter cell type as microglia was confirmed by morphologic features, tissue distribution, immunoelectron microscopy, and double staining with monoclonal antibodies or lectins to macrophage markers, including class II major histocompatibility antigens. The two cell types had different disposition in senile plaques of elderly individuals and of those with Alzheimer's disease. Astrocytes were present at the periphery of the plaques, whereas microglial cells were centrally placed, often in juxtaposition to amyloid. The results are discussed with respect to ontogeny of glial cells and the ability of monoclonal antibodies to recognize epitopes on unrelated proteins.

Antibodies, Monoclonal↗
PubMed1996

Autometallographic localization of protein-bound copper and zinc in the common winkle, Littorina littorea: a light microscopical study.

Copper (Cu), zinc (Zn) and calcium (Ca) were demonstrated histochemically by means of conventional stains (rubeanic acid for copper, dithizone for zinc, and cobalt nitrate for calcium) and by autometallography in various tissues of winkles (Littorina littorea) sublethally exposed to either copper or zinc dissolved in sea water. Rubeanic acid and dithizone procedures exhibited poor sensitivity: there was no positive reaction after fixation tissues with Bouin's fixative, and only a weak reaction after ethanol fixation. Autometallography, however, produced a positive reaction with both fixatives in the form of black silver deposits in some key cell types. In winkles not exposed to either copper nor zinc, autometallographically demonstrated metals were found in the connective tissue pore cells, the lysosomes of digestive cells, the basal lamina of the digestive tubule epithelium, and cytoplasmic granules in the epithelial cells of the stomach wall. In addition, in winkles exposed to copper, metal deposits were present in some apical cytoplasmic granules of ciliated cells in the gill epithelium, the mucous secretion of gill mucocytes, and the circulating haemocytes. In winkles exposed to zinc, metal deposits were found in the basal cytoplasmic granules of ciliated cells in the gill epithelium, the mucous secretion of gill mucocytes, the apex and basal lamina of the nephrocytes in the kidney, and the connective tissue layer surrounding the blood vessels. Additionally, calcium was demonstrated histochemically in the cytoplasm of digestive cells, the cytoplasm of the epithelial cells of the stomach wall, the mucocytes of gills, the basal lamina of the kidneys, the haemocytes, the calcium and pore cells of connective tissue, and the oocyte cytoplasm. Metals were not detected by any procedure in sperm cells, in the cytoplasmic granules of oocytes, or in the basophilic cells in the digestive tubules. In conclusion, autometallography is a highly sensitive method and provides an excellent tool to localize protein-bound copper and zinc in molluscan tissues, and its use in combination with conventional histochemical or chemical methods is highly recommended.

Animals↗
PubMed1991

A novel method for optimum biopsy specimen preservation for histochemical and immunohistochemical analysis.

A novel method has been developed for optimally processing biopsy specimens combining freeze-substitution with low-temperature plastic embedding. Immunohistochemistry and conventional histochemical stains were all readily performed on tissue displaying high-quality morphologic preservation. Labile antigens, especially lymphoid cell surface antigens, were well preserved. This new method avoids the need for tissue fixation and combines the superior morphologic preservation of fixed embedded tissue with the reactivity of cryostat sections. This method ensures that diagnostic information from even the smallest biopsy specimen is maximized because a wide range of phenotypic markers can be applied and evaluated in relation to high-quality morphologic preservation of tissue. Biopsy specimens are stored at room temperature without loss of tissue-specific characteristics during storage.

Acetone↗
PubMed1985

[Use of semithin sections for the histochemical study of carbohydrate-containing biopolymers in cells and tissues with the aid of lectins].

A sensitive method of histochemical analysis of cellular and tissue glycoconjugates on semithin sections using lectins is suggested. For fixation tissue bioptates were incubated for 4 h in a 2.5% glutaraldehyde in phosphate buffered saline (PBS) at 4 degrees C, then washed for 1 h in 0.2 M glycine in PBS. After epon-araldite embedment and preparation of semithin sections, the resin was removed in saturated ethanol-KOH solution during 5-10 s. Endogenous perooxidase was inactivated in methanol containing 0.3% H2O2. For identification of lectin-binding sites semithin sections were incubated for 30 min in a 0.005% solution of lectin-peroxidase conjugate in PBS and visualized by 0.05% diaminobezidine solution in PBS, containing 0.015% H2O2. The method described ensures good preservation of cellular and tissue glycoconjugates and is highly specific and sensitive.

Animals↗
PubMed1988

Detection of cell proliferation in pig testis and intestine sections using monoclonal anti-bromodeoxyuridine antibody and immunogold silver staining.

For the first time a monoclonal antibody against 5-bromodeoxyuridine was used to detect cell proliferation in pig testis and intestine sections. The influence of several parameters such as mode of injection, addition of thymidine biosynthesis inhibitor, tissue fixation, hydrolysis and revelation was examined. The technique of choice consisted in intravenously injecting the animals with 50 mg/kg BUdR added to 10 mg/kg FUdR 2 h before tissue collection and Bouin fixation; hydrolysis of sections was performed by HC1 4N: Ethanol 70 degrees (1:1 v/v); revelation of BUdR was made by a secondary antibody linked to colloidal gold particles, followed by a silver enhancement step. The data were superior when compared to those obtained by direct immunofluorescence and by the PAP technique. The described method is convenient and sensitive, provides an intense nuclear labelling without background and allows simultaneous examination of histology. The advantages over the technique using tritiated thymidine are particularly obvious when fast screening of numerous samples is required or when new experimental protocols are developing.

Animals↗
PubMed2002

A precise radiographic technique for the measurement of dimensional changes in heart valve biomaterials following fixation.

Accurate tissue thickness measurements are difficult to acquire by present techniques. Error is introduced by tissue compression during measurements or by tissue processing prior to measurement. In the field of valve replacement, tissue dimensional changes from fixation prior to implantation may predispose implants to premature tissue failure and it becomes important to have an accurate method for comparing cusp dimensions pre- and post-fixation. A new approach is to use high-resolution digital radiography to make thickness maps of entire specimens. For 25 matched porcine aortic valve cusps, we have evaluated this technique's ability to measure and compare thickness, surface area and volume before and after 7 days of aldehyde fixation. Digital radiographs were acquired pre- and post-0.5% glutaraldehyde (n=13) or 10% formaldehyde (n=12) fixation. Mean thickness, surface area, volume and four measurements to evaluate shape differences with fixation were obtained and compared pre- and post-fixation using paired t tests. The results demonstrate that this X-ray imaging technique can provide dimensions of matched fresh and fixed specimens and is sensitive enough to show statistically significant changes due to fixation. These findings also illustrate that aldehyde fixation can cause tissue contraction resulting in a significant overall increase in tissue thickness and a decrease in surface area. This technique could be used to gain further insights into tissue anatomy and mechanics.

Animals↗
PubMed2004

Allograft interference screw fixation in meniscus transplantation.

Allograft meniscus transplantation is indicated to restore proper knee biomechanics and prevent subsequent articular degeneration in patients with a meniscus-deficient knee. A variety of techniques for fixation of meniscal transplants exist, with some techniques using soft-tissue fixation of the meniscal horns and others using bony fixation. The authors present a technique of meniscus transplantation using a tibial slot with allograft interference screw fixation. We have had excellent early results with this technique and believe that a cortical allograft interference screw reliably maintains the anatomic position of the meniscal attachments that are critical to the biomechanics and functional outcome of meniscus transplantation.

Arthroscopy↗
PubMed1978

The Golgi rapid method in clinical neuropathology: the morphologic consequences of suboptimal fixation.

Cytologic changes in neurons of the neocortex of mice consequent to suboptimal fixation have been investigated systematically in Golgi-rapid preparations. With few exceptions, there is no alteration in cellular morphology if the brain is refrigerated after death, and fixed by immersion within 3 hours. With latencies of fixation of 6 hours or more, autolytic changes supervene which modify the general histologic appearance and the morphology of individual cells. In general, the degree of tissue and cellular change is proportional to the latency between death and tissue fixation. Similar alterations in cellular morphology and general tissue appearance are found in Golgi-rapid impregnations of human brains obtained at autopsy. However, the degree of tissue autolysis in the human specimens bears a less predictable relationship to the latency of fixation after death. The duration of preterminal metabolic encephalopathy appears to be equally decisive as a determinant of tissue preservation.

Adolescent↗
PubMed2001

Chloride distribution in the CA1 region of newborn and adult hippocampus by light microscopic histochemistry.

GABA, the main inhibitory neurotransmitter in the central nervous system, exerts its effect by rendering the postsynaptic GABAA receptors permeable to chloride ions. Thus, depolarizing or excitatory effects of GABA, experienced in early postnatal life or in certain regions and/or conditions of the adult brain, is thought to be associated with a reversed transmembrane chloride gradient. However, there is only limited direct information about the correlation of the actual excitatory versus inhibitory effects of GABA and the local chloride distribution. Precipitation of chloride with silver is a potential way to immobilize and visualize chloride ions in biological tissue. We examined the applicability of light microscopic histochemistry, based on trapping tissue chloride with silver ions during freeze-substitution or aldehyde fixation, to visualize the chloride distribution in hippocampal slices. The freeze-substitution procedure yielded better chloride retention while with aldehyde fixation tissue preservation was more appropriate. Both methods were qualitative only, had limited applicability to the superficial 20-30 microns of slices, but were able to demonstrate a reduced extracellular-to-intracellular chloride gradient in the CA1 pyramidal neurons of the newborn hippocampus as compared to adult animals. In the 4-aminopyridine model of epilepsy, redistribution of chloride from extracellular to intracellular space could also be demonstrated.

4-Aminopyridine↗