Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: KEGG:D02003 (NBT)
 1,323 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

To avoid extensive manipulation for the purification of RNA from cells, several methods were evaluated for the direct release of RNA from influenza virus infected cells and supernatants using slot blot hybridization and non-radioactive probes. Treatment with an equal volume of 10 M aqueous guanidine hydrochloride produced the best hybridization signal. Less, but significant amounts of RNA were also released using the following treatments: dilute alkali (final concentration of 0.16 M NaOH) or 100 degrees C/5 min or RNA sample buffer containing formamide/formaldehyde, then heating at 65 degrees C/10 min. Despite the presence of large amounts of cell debris, RNA from guanidine hydrochloride treated whole cell extracts bound quantitatively to the positively charged nylon membranes. The sensitivity of RNA detection when whole cell extracts treated with guanidine hydrochloride were probed with a digoxigenin labelled cDNA probe was similar to the detection of RNA in highly purified, protein free samples. Three positively charged membranes were tested (from Amersham, ICN and Boehringer Mannheim) using two alkaline phosphatase substrates, NBT-X phos, and a chemiluminescent substrate, 3-(2'-spiroadamantane)-4-methoxy-4-(3'-phosphoyloxy)-phenyl- 1-1,2-dioextane (AMPPD) and a peroxidase substrate, tetramethylbenzidine (TMB). The Boehringer Mannheim membrane had the highest sensitivity for the alkaline phosphatase substrates, but the peroxidase reaction with the TMB substrate was the most consistently sensitive, irrespective of which membrane was used. The ability to quantitatively detect RNA from whole cells without any purification will allow the rapid screening of large numbers of samples for specific RNA species in research or diagnostic laboratories.
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PMID:Rapid treatment of whole cells and RNA viruses for analysis of RNA by slot blot hybridization. 162 16

A complete evaluation of neutrophil function including: chemotaxis; adhesion; aggregation; phagocytosis; granule content and degranulation; respiratory burst activity; and bacterial killing; is expensive and requires the services of a specialized laboratory. However, preliminary screening of a patient with a predisposition toward infection, can be carried out using simple and inexpensive methods. These include examination of blood films, which may prove helpful in the diagnosis of Chediak-Higashi syndrome and specific granule deficiency; the Rebuck skin window test, which estimates chemotactic defects; the NBT test, which screens for chronic granulomatous disease patients; and peroxidase staining of the blood film in order to estimate the content of myeloperoxidase, when myeloperoxidase deficiency is suspected. For final diagnosis and determination of genetic transmission and radical treatment, ie, bone marrow transplantation, specific tests are indicated. Neutrophil function studies have also proved useful in detecting diseases in which defects in neutrophil function are secondary to the primary disorder. Indeed, increased neutrophil chemotaxis has been reported in the active phase of diseases such as: familial Mediterranean fever; psoriasis vulgaris, Behcet's syndrome and Sweet's syndrome. In these disorders the neutrophil chemotaxis assay has aided in the diagnosis and follow-up, particularly in evaluating the response to antiinflammatory agents, such as colchicine.
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PMID:Neutrophil function studies in clinical medicine. 298 Feb 76

Basophilic granulocytes were purified from the blood of normal individuals by successive isopyknic centrifugation and elutriation centrifugation. Starting with the leukocyte-rich fraction of 500 ml of blood, we recovered 31 to 80% (mean 51%, n = 20) of the basophils in 45 to 87% purity (mean 69%, n = 23). The contaminating cells were mainly lymphocytes. The basophils were greater than 98% vital (exclusion of ethidium bromide and hydrolysis of fluorescein diacetate). The histamine content of the basophils was 1.1 to 2 pg/cell (mean 1.6 pg/cell, n = 22). With anti-IgE, 30 to 50% of the histamine was released; with phorbol myristic acetate (PMA) or the calcium ionophore A23187, 70 to 100% of the histamine was released. Serum-opsonized zymosan (STZ) did not induce histamine release. Reactions with monoclonal antibodies revealed that the basophils expressed the C3bi receptor (CR3) and the leukocyte function-associated antigen 1 (LFA1), but not the gp 150,95 antigen, the C3b receptor (CR1), or the low avidity Fc gamma receptor. Basophils carry class I but not class II HLA antigens. During incubation of the basophils with serum-opsonized Staphylococcus aureus or Escherichia coli, these bacteria were neither phagocytized nor killed. STZ, PMA, A23187, or anti-IgE did not initiate an "oxidative burst" in the basophils. This was tested with oxygen consumption, cytochrome c reduction, NBT reduction, chemiluminescence, and release of hydrogen peroxide. Moreover, we did not detect cytochrome b558, superoxide dismutase, catalase, or peroxidase in the basophils. Of the typical granule-associated enzymes lysozyme, Vitamin B12-binding protein, and beta-glucuronidase, only beta-glucuronidase was present in the basophils in detectable amounts. This enzyme was released, together with histamine, on incubation of the cells with PMA, A23187, or anti-IgE, but not with STZ. We conclude that basophils from normal human blood are not phagocytes and are probably not involved in the oxidative defense of the host against foreign antigens.
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PMID:Metabolic comparison between basophils and other leukocytes from human blood. 300 19

A study was carried out to determine the effect of protein deficiency on the phagocytic function of blood neutrophils and of peritoneal exudate of rats. The deficient animals exhibited significantly lower leukocyte and neutrophil values, as well as NBT reduction and diminished peroxidase and bactericidal capacity. Englobement of S. aureus and latex particles was found to be normal in both groups. Alkaline phosphatase activity in the neutrophils appear to be increased in the deficient animals.
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PMID:[Influence of protein malnutrition on the phagocytic function of neutrophils in rats]. 343 16

A myeloid leukemia cell line (TK-1) was established from the peripheral blood of a patient with lymphoblastic lymphoma whose leukemia cells were composed of T-lymphoblasts and immature myeloid cells. The established TK-1 cell line consisted of immature myeloid cells with heavy azurophilic granulation in the cytoplasm. The TK-1 cells were positive for peroxidase, and exhibited a strong positive reaction for alpha-naphthyl acetate esterase and naphthol AS-D chloroacetate esterase. The cells were weakly positive for Fc gamma-receptors, showed no phagocytosis and did not reduce NBT. With the treatment of 1 alpha, 25-dihydroxy-vitamin D3, they exhibited morphological and functional differentiation. The TK-1 cell line contained normal diploid cells and pseudodiploid cells, and the two populations were successfully cloned; the clone with a normal karyotype was designated the TK-1D cell line, and the clone with a pseudodiploid karyotype, which had a translocation involving chromosomes 14, 17 and one other chromosome, was designated the TK-1B cell line. These cloned cells lacked Epstein-Barr virus nuclear antigens and had almost the same myelomonocytic characteristics as the parent TK-1 cells. The breakpoint of chromosome 17 involved in the translocation of the pseudodiploid cells was identified to be a band 17q23.
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PMID:Establishment of a new myeloid leukemia cell line (TK-1), and isolation of cells having a translocation involving a band 17q23. 345 71

A number of immunocytochemical detection systems for determining the chromosomal localization of specific nucleic acid sequences by non-radioactive in situ hybridization have been compared. The procedures were: 1. the peroxidase/diaminobenzidine (PO/DAB) combination, either or not gold/silver intensificated; 2. alkaline phosphatase marking using the nitro-blue tetrazolium plus bromochloro-indolyl phosphate substrate combination (AP/NBT + BCIP); and 3. immunogold with or without silver enhancement. The procedures were first tested and optimized in dot blot experiments and then applied to in situ hybridization. As hybridization probes, both a middle-repetitive and a unique sequence (modified with 2-acetylaminofluorene (AAF] were used. The advantages and disadvantages of the various methods for reflection contrast (RC) or transmission electron microscopic (TEM) visualization of hybrids are discussed.
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PMID:Non-radioactive in situ hybridization. A comparison of several immunocytochemical detection systems using reflection-contrast and electron microscopy. 361 Jun 73

A new method for the determination of xanthine oxidase activity, based on the oxidation of 2,2'-azino-di(3-ethylbenzthiazoline-6-sulphonate) (ABTS) by use of uricase and peroxidase, is described. The absorbance increase of the oxidized form of ABTS, measured after 10 min at 410 nm is proportional to xanthine oxidase activity. The method is sensitive, precise (CV below 8.3%), and linear up to 20 U/l. The analytical recovery of the ABTS-method was quantitative. Comparison with the UV and colorimetric NBT-method gave good correlation (r greater than or equal to 0.984). Reference values for serum xanthine oxidase activities determined with the new ABTS-method on 83 healthy persons are 0 to 1.20 U/l.
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PMID:Spectrophotometric assay of xanthine oxidase with 2,2'-azino-di(3-ethylbenzthiazoline-6-sulphonate) (ABTS) as chromogen. 380 44

Multinucleated giant cells of mononuclear phagocyte origin (monocyte or macrophage polykaryons [MPs] ) are seen in numerous different normal and pathologic states. We have previously shown that gamma interferon (IFN-gamma) induces fusion of uninuclear monocytes (UMs) to form MPs. This study was designed to characterize these IFN-gamma-induced MPs. Control and IFN-gamma-treated UMs and MPs did not have peroxidase activity, but they stained intensely for nonspecific esterase and acid phosphatase. The esterase of UMs and MPs was abolished by fluoride, but the acid phosphatase of UMs and MPs was only minimally decreased by tartrate. The phagocytosis of polystyrene spheres and glutaraldehyde-fixed erythrocytes by MPs was moderately depressed as compared with control or treated UMs, whereas the phagocytosis of IgG-coated erythrocytes was markedly depressed. Populations of control monocytes produced less H2O2 in response to 200 nmol/L of phorbol myristate acetate than did IFN-gamma-treated monocytes (37 +/- 7 v 199 +/- 29 nmol/h per milligram of cell protein). However, when examined microscopically, individual MPs had less ability to reduce NBT (18% +/- 5% positive for MP, 91% +/- 3% for treated UMs, and 67% +/- 3% for control UMs). The surface membrane antigens Leu M3, OKM1 (C3bi receptor), DU-HL60-3, DU-HL60-4, TE5, and V1 were not expressed or were expressed poorly in MPs; they were expressed normally in control and treated UMs. However, HLA-DR expression was increased in treated UMs and MPs. The binding of the lectins RCA, Con A, WGA, DBA, UEA, and PNA was equivalent in all cells. Thus, MPs formed by fusion of UMs in vitro after culture with IFN-gamma differ in several features from UMs.
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PMID:Phenotypic characterization of gamma interferon-induced human monocyte polykaryons. 393 91

In thyroid gland, iodination takes place on the apical plasma membrane and requires the presence of the thyroid peroxidase and H2O2 generating system. H2O2 generation and NBT (nitro blue tetrazolium) reductase activity (both of which are NADPH-dependent) as well as peroxidase activity were compared for their respective orientations in membrane vesicles. The possible role of NADPH-NBT reductase activity in H2O2 generation was also examined. Results favor the conclusion that thyroid peroxidase is oriented towards the luminal side of the vesicles, whereas the NADPH site of NADPH oxidase-dependent H2O2 generation is located on the external side of the same or of different vesicles. Furthermore, it is shown that different NADPH-NBT reductase activities are present on both the outer and inner surfaces of the membrane vesicles, and that none of these activities is able to produce either H2O2 or O-2. The idea that a multi-component complex is involved in H2O2 generation is discussed, and a model is proposed which takes into account the possible spatial separation of the thyroid peroxidase site from the NADPH site of this H2O2 generation system on the apical membrane of the thyrocyte.
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PMID:Relation between thyroid peroxidase, H2O2 generating system and NADPH-dependent reductase activities in thyroid particulate fractions. 401 97

In 106 workers (47 women and 59 men) being in professional contact with organic solvents containing benzene and its homologues during 1 to 122 months the cytochemical examination of peripheral blood neutrophils has been performed. The patterns of neutrophil functional activation have been noted expressed in increased activities of acid phosphatase and beta-glucuronidase, increased NBT reduction and diminished glycogen reserves. Those changes were accompanied by diminished peroxidase and alkaline phosphatase activities. The stimulated NBT reduction, elevated in majority of workers, exhibited negative correlation with the exposure time what indicates the practical value of that test monitoring the biological effects of professional contact with the solvents.
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PMID:Cytoenzymatic studies on neutrophils in workers having contact with organic solvents containing benzene, toluene and xylene. 616 42


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