Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.3.1 (alkaline phosphatase)
47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Since our previous experiments suggested that glycosylation-inhibiting factor (GIF) is a phosphorylated derivative of a phospholipase inhibitory protein, we determined whether other well-known phospholipase inhibitors may have similar biological activities. The results showed that phospholipase A2 (PLA2) inhibitors, such as recombinant human lipocortin I and ONO-RS-082, could switch T cell hybridoma 12H5 cells from the formation of glycosylated IgE-binding factors (IgE-BF) to the formation of unglycosylated IgE-BF, whereas neomycin, a phospholipase C inhibitor, failed to affect the nature of IgE-BF formed by the cells. The minimum concentrations of lipocortin I and ONO-RS-082 required for switching the 12H5 cells to the formation of unglycosylated IgE-BF were comparable to or less than IC50 of the inhibitors for PLA2. The ability of partially purified GIF to switch the 12H5 cells to the formation of unglycosylated IgE-BF was markedly enhanced by treatment of the preparation with alkaline phosphatase. It was also found that lipocortin I and ONO-RS-082, but not neomycin, facilitated the generation of GIF-producing T cells. When spleen cells of ovalbumin (OVA)-primed BDF1 mice were stimulated with homologous antigen and the activated T cells were propagated by recombinant IL-2 in the presence of GIF, lipocortin I, or ONO-RS-082, T cells obtained in the cultures constitutively produced their own GIF. Antigenic stimulation of the T cells induced the formation of unglycosylated IgE-BF and GIF with an affinity for OVA.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Effect of phospholipase A2 inhibitors on mouse T lymphocytes. I. Phospholipase A2 inhibitors exert similar immunological activities as glycosylation inhibiting factor. 253 36

Existing methods of measuring IgE in in vitro peripheral blood lymphocyte (PBL) cultures are not sufficiently sensitive to detect IgE when it is present in small amounts. This paper describes a modification of a two-site ELISA which increases the sensitivity of the assay 10-20-fold. By using the Fab' fragment of either rabbit or mouse monoclonal anti-IgE conjugated to alkaline phosphatase (AP) as the detector, the background of the assay was reduced sufficiently to permit signal amplification, using a commercially available amplified AP substrate. With this assay as little as 10 pg/ml of IgE could be detected. The interassay coefficient of variation was 15-18% between 1200 and 100 pg/ml IgE (n = 14) and there was a good correlation with a commercial IgE radioimmunoassay (RIA) (r = 0.98, n = 38).
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PMID:Ultrasensitive enzyme-linked immunosorbent assay (ELISA) for the detection of picogram quantities of IgE. 273 14

A modified ELISA was developed to estimate antigen-specific IgE antibody. IgE against ragweed antigen E (AgE) was the model system, and paired pre- and postimmunotherapy sera were evaluated because the latter had been demonstrated to contain significant amounts of IgG antibody. Patients' serum IgE was bound to the surface of wells of a polystyrene microtiter plate previously coated with monoclonal murine antihuman IgE. After washing, AgE conjugated with alkaline phosphatase was added and was specifically bound by IgE. Optical density was recorded after addition of enzyme substrate to the wells and was proportional to the concentration of IgE directed against ragweed AgE in the serum samples. In contrast to many other assay techniques for antigen-specific IgE, interference by specific antibody of other classes cannot occur with this method. Results correlated well with those of a previously described solid-phase radioimmunoassay that required myeloma IgE.
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PMID:Estimation of IgE antibody with a nonisotopic technique with no interference of isotypic antibodies. 319 64

In this paper we outline a flexible and rapid method to measure picogram quantities of isotype-specific immunoglobulin (Ig), including IgE. Only readily or commercially available reagents are required: isotype-specific, anti-human Ig murine monoclonal antibodies (Mab) to coat microtiter plates, polyclonal alkaline phosphatase-coupled isotype-specific F(ab)'2 or Fab' fragments as second antibodies, and an enhanced developing system that amplifies the signal-to-noise ratio of the quantitatively bound second antibody. The procedure is detailed in the appendix to enable easy application, even if one has no previous experience with ELISAs. This system can be used to detect less than 10 picograms of Ig in cultures supernatants of cells that contain mixtures of various Igs and it can be used to detect the product of a single cell producing Ig. This method also will be applicable to measurement of the minute quantities of lymphokines and other biologically active molecules produced in vitro and found in various fluids in vivo.
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PMID:Enhanced ELISA: how to measure less than 10 picograms of a specific protein (immunoglobulin) in less than 8 hours. 326 91

During the period from 1982 to 1985, biosynthetic growth hormone (methionyl-hGH) was administered to 55 children with pituitary growth deficiency, 49 with idiopathic, six with other forms of the disorder. Preparations with a relatively high content of Escherichia coli polypeptides (ECP) were given to 32 children (group I), those with a markedly reduced ECP content to 12 (group II), while 11 children received almost ECP-free preparations (group III). The treatment achieved an intensive rise of growth acceleration in all three groups. At the same time, somatomedin C, biological somatomedin activity and alkaline phosphatase rose steeply, reaching a maximum after six months and remaining at high concentrations even thereafter. High antibody titres against ECP and hGH (with high binding capacity) were demonstrated in group I children, significantly lower titres with little binding capacity in those of group II, while most of those in group III had no measurable antibodies. Even in high concentrations the antibodies did not have any inhibiting effect on growth. No positive correlation between antibody titre and growth velocity was demonstrable. An exception was a boy with an allergy in group I, who had a maximal antibody titre and, at the same time, a high serum concentration of IgE. He had marked delay in growth which, however, quickly became normal on administration of extractive hGH.
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PMID:[Therapy of pituitary dwarfism with recombinant human growth hormone. A multicenter study]. 351 93

Mouse bone marrow-derived mast cells passively sensitized with monoclonal IgE released paf-acether (platelet-activating factor) and beta-hexosaminidase when challenged with the specific antigen. The formation and the release of paf-acether followed an early increase in the activity of the acetyltransferase, the main enzyme in paf-acether biosynthesis. The antigen-induced activation of the acetyltransferase was dependent on physiologic temperature and on the presence of Ca2+. By using microsomal fractions from unchallenged and challenged mast cells, the Vmax values were 3.5 and 12.0 nmol/min/mg of protein, respectively, whereas in both cases a Km value for acetyl-coenzyme A of 172 microM was measured. The stimulation of acetyltransferase could be mimicked in vitro under experimental conditions which favor phosphorylation, i.e. adding ATP and Mg2+ to lysates from unchallenged mast cells. In contrast, ATP and Mg2+ were uneffective on lysates from challenged cells that exhibited high level of acetyltransferase activity, suggesting that phosphorylation of the enzyme already took place at the time of cell stimulation. Moreover, addition of alkaline phosphatase to microsomal fraction obtained from either antigen-challenged mouse bone marrow-derived mast cells or unchallenged cells, resulted in 52% and 43% loss of acetyltransferase activity, respectively. Phorbol myristate acetate treatment of cells doubled the enzyme activity supporting the phosphorylation hypothesis. Thus, we report on the immunologic activation of a key enzyme for paf-acether synthesis and on the mechanism of this activation in a pure mast cell population. A link between bridging of IgE receptors and the activation of an enzyme critical to the formation of a lipid mediator is thereby evidenced.
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PMID:Biosynthesis of paf-acether. IX. Role for a phosphorylation-dependent activation of acetyltransferase in antigen-stimulated mouse mast cells. 358 83

IgE RF was measured by ELISA assay using aggregated IgG as a solid phase immunosorbent and alkaline phosphatase-conjugated Fc epsilon-specific monoclonal and polyclonal antibodies as indicators. The presence of IgE RF was defined in this assay as binding of the conjugate greater than 2.33 SD above the mean for control sera (N = 27). Total IgE was elevated in 25% (13/52) of sera of patients with seropositive Rheumatoid Arthritis (RA), yet was normal in the sera of 17/19 Mixed Cryoglobulinemia (MC) patients. IgE RF was present in 33% (21/63; p less than 0.05) RA sera, and none of sera from 19 MC patients tested. It did not correlate with IgM RF titer or total IgE, and was not detected in separated IgM and IgG fractions of 7 purified mixed cryoglobulins from patients with MC. These findings suggest that IgE RF may not be an important pathogenic factor in the clinical manifestations of MC. Its potential significance in RA is discussed.
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PMID:Prevalence of IgE rheumatoid factor (IgE RF) in mixed cryoglobulinemia and rheumatoid arthritis. 379 15

The use of antibody to link antigen to microtitre plates in the enzyme-linked immunosorbent assay (ELISA) has been extended to include mouse monoclonal antibodies and polyspecific rabbit antibodies. Small amounts of both reagents could be used. The capacity of the microtitre plate for the antibody was found to be critical and irradiated plates generally gave better results. Both rabbit anti-IgE conjugated to horseradish peroxidase, and monoclonal mouse anti-IgE with alkaline phosphatase-conjugated rabbit anti-mouse IgG could be used as detection reagents. Comparison with the radioallergosorbent (RAST) test showed a good agreement with the sandwich ELISA. The sandwich ELISA using polyspecific rabbit antibody was substantially more sensitive than conventional ELISA and also marginally more sensitive than RAST.
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PMID:The use of monoclonal and polyspecific antibodies in the IgE sandwich ELISA. 395 Apr 24

A new system is described for the enumeration of human immunoglobulin-secreting cells (ISC), based upon the ELISA methodology. In principle, putative ISC are incubated over a solid phase containing bound anti-Ig of the isotype being tested. Secreted Ig is immobilized at or near the point of release from the ISC, and the resulting Ig fingerprint of the ISC is then visualized by the sequential application of an anti-Ig-alkaline phosphatase conjugate, followed by a substrate-agarose overlay. The system is capable of detecting IgE-secreting cells, and pokeweed mitogen-stimulated IgG-secreting cells with sensitivity at least equivalent to the protein A hemolytic plaque assay.
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PMID:Enumeration of human immunoglobulin-secreting cells by the ELISA-plaque method: IgE and IgG isotypes. 636 82

Splenic T lymphocytes of rats treated with complete Freund's adjuvant (CFA) spontaneously release a lymphokine that inhibits the glycosylation of IgE-binding factors during their biosynthesis and provides the factors with biologic activity: selective suppression of the IgE response. The lymphokine, which is called glycosylation-inhibiting factor, also prevents IgE-induced increases in Fc epsilon R+ cells. The glycosylation inhibiting factor is formed by stimulation of BCG-primed spleen cells with PPD and participates in the selective formation of IgE-suppressive factors. The lymphokine is derived from OX 8+ T cells in both the CFA and BCG systems. The glycosylation-inhibiting factor is a 16,000-dalton peptide, as estimated by gel filtration, and specifically binds to monoclonal antibody against lipomodulin, a phospholipase inhibitory protein. Furthermore, "lipomodulin" was detected by radioimmunoassay in the 16,000-dalton fraction that contained glycosylation inhibiting factor. The fraction did not inhibit phospholipase A2 but after alkaline phosphatase treatment, the fraction did inhibit phospholipase A2. Furthermore, purified lipomodulin obtained from glucocorticoid-treated rabbit neutrophils had the same biologic activities as glycosylation inhibiting factors; i.e., it inhibited both protein glycosylation of IgE-binding factors and IgE-induced expression of Fc epsilon R. The results collectively indicate that glycosylation-inhibiting factor is a fragment of phosphorylated lipomodulin.
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PMID:Modulation of the biologic activities of IgE-binding factors. I. Identification of glycosylation-inhibiting factor as a fragment of lipomodulin. 660 Feb 57


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