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)

Ubiquinol-1 in aerated aqueous solution inactivates several enzymes--alanine aminotransferase, alkaline phosphatase, Na+/K(+)-ATPase, creatine kinase and glutamine synthetase--but not isocitrate dehydrogenase and malate dehydrogenase. Ubiquinone-1 and/or H2O2 do not affect the activity of alkaline phosphatase and glutamine synthetase chosen as model enzymes. Dioxygen and transition metal ions, even if in trace amounts, are essential for the enzyme inactivation, which indeed does not occur under argon atmosphere or in the presence of metal chelators. Supplementation with redox-active metal ions (Fe3+ or Cu2+), moreover, potentiates alkaline phosphatase inactivation. Since catalase and peroxidase protect while superoxide dismutase does not, hydrogen peroxide rather than superoxide anion seems to be involved in the inactivation mechanism through which oxygen active species (hydroxyl radical or any other equivalent species) are produced via a modified Haber-Weiss cycle, triggered by metal-catalyzed oxidation of ubiquinol-1. The lack of efficiency of radical scavengers and the almost complete protection afforded by enzyme substrates and metal cofactors indicate a 'site-specific' radical attack as responsible for the oxidative damage.
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PMID:Enzyme inactivation by metal-catalyzed oxidation of coenzyme Q1. 135 46

Progesterone (P)-induced PRL secretion in estradiol (E)-primed monkeys is not due to direct pituitary stimulation, because lactotropes do not express progestin receptors (PR). However, the hypothalamus, particularly the tuberoinfundibular dopaminergic system (TIDA), plays a major role in the regulation of PRL secretion. To determine whether hypothalamic dopamine neurons are progestin target cells, the colocalization of PR and tyrosine hydroxylase (TH), a phenotypic marker of dopaminergic neurons, was examined with double immunocytochemistry. Two methods for visualizing the antigens were applied; the first was a dual peroxidase method, and the second was a peroxidase-alkaline phosphatase method. In addition, the question of whether E induces PR in dopamine neurons was explored. Spayed female monkeys were treated with empty Silastic capsules, E-filled capsules for a period of 28 days, or E capsules supplemented with P capsules for the last 14 days of E treatment. Only the E- plus P-treated monkeys exhibited an increase in serum PRL during the P treatment period. Frontal sections at the level of the optic chiasm and arcuate nucleus were examined for the colocalization of TH and PR. After E treatment, hypothalamic PR-positive cells increased in both intensity and number. Neurons expressing both TH and PR were detected in the rostral hypothalamus, lateral to the third ventricle (A11-rostral) and in a discrete subventricular population (A11-subvent). The lateral population continued caudally (A11-caudal). The A11-subvent population exhibited little steroid regulation. Of the remaining A11 TH neurons, approximately 20% exhibited PR in the spayed and E-treated groups. Addition of P doubled the percentage of PR-containing TH neurons in this group. Although very few TH-positive neurons in the ventral arcuate nucleus contained PR (A12-ventral), many double labeled neurons were observed in the dorsal arcuate region (A12-dorsal). Ventral arcuate TIDA neurons were not regulated by steroids, but E plus P increased PR expression in A12-dorsal. Double labeled cells were rarely seen in the zona incerta (A13) or the emerging ventral tegmental area (A10). In summary, P probably does not act directly on ventral arcuate TIDA neurons to stimulate PRL secretion. However, the frequency of PR-positive dopamine neurons in the A11-rostral, A11-caudal, and A12-dorsal groups increased with E and P treatment. Therefore, the contribution of the PR-positive periventricular dopamine neurons to progestin-stimulated PRL secretion may be important.
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PMID:Immunocytochemical colocalization of hypothalamic progestin receptors and tyrosine hydroxylase in steroid-treated monkeys. 135 39

In order to investigate the possible involvement of corticotropin-releasing factor (CRF) and somatostatin (SRIF) on thyrotropin-releasing hormone (TRH) neuronal cell activity in the rat hypothalamic paraventricular nucleus, we have proceeded to the simultaneous localization of CRF or SRIF and TRH. For this purpose, we used a dual immunostaining procedure that employed antibodies to CRF and SRIF and peroxidase-labeled goat anti-rabbit IgG as a first sequence, and antibodies to a cryptic fragment (Phe178-Glu199) of pro-TRH (to label TRH neurons) and alkaline phosphatase-labeled goat anti-rabbit IgG as the second sequence. A rich innervation of the paraventricular nucleus by immunoreactive CRF and SRIF fibers was observed. A large number of CRF and SRIF nerve endings were seen intimate anatomic proximity and often appeared to surround TRH-containing cell bodies. These results strongly suggest that TRH neurons might be regulated by both CRF and SRIF. These interactions might be the neuroanatomical basis for the already observed inhibitory effects of CRF and SRIF on TRH release.
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PMID:Neuroanatomical connections between corticotropin-releasing factor (CRF) and somatostatin (SRIF) nerve endings and thyrotropin-releasing hormone (TRH) neurons in the paraventricular nucleus of rat hypothalamus. 135 8

A simple to use, robust, quantitative, and extremely sensitive colorimetric assay for alkaline phosphatase (EC 3.1.3.1), designed to be used as a detection system in diagnostic assays employing antibodies or gene probes, is described. This technology is based on the novel principle of prosthetogenesis, according to which a purpose-designed substrate (a prosthetogen) for a primary analyte-linked enzyme label is hydrolyzed to produce a prosthetic group for a detector enzyme system. The prosthetogen employed here is a derivative of FAD which is phosphorylated at the 3'-position of the ribose ring (FADP), the label enzyme is alkaline phosphatase, and the detector is a D-amino-acid oxidase/horseradish peroxidase-coupled system. Essentially each turnover of every molecule of alkaline phosphatase produces a molecule of D-amino-acid oxidase for detection. Thus enormous amplification of the initial signal is achieved in short time periods because of the relatively high turnover number of alkaline phosphatase for FADP. The system can be formatted as a stable, preformed, freeze-dried preparation containing all analytical components, which is reconstituted simply by addition of buffer solution. This methodology can quantitate less than 0.1 amol of alkaline phosphatase in 30 min at 25 degrees C using microtiter plates.
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PMID:Amplified assay of alkaline phosphatase using flavin-adenine dinucleotide phosphate as substrate. 136 Jul 71

A novel methodology for coupling liquid-liquid extraction with affinity interaction has been developed to selectively and efficiently purify and separate glycoproteins. The basis for the separation is the selective extraction of glycoproteins from an aqueous solution into a reverse micellar organic phase by using concanavalin A (a sugar-binding lectin) as a facilitative carrier. Specifically, horseradish peroxidase (a common glycoprotein) can be bound to concanavalin A in an aqueous phase and then extracted into an AOT-isooctane organic phase with negligible loss in enzyme activity. Virtually no extraction of peroxidase occurs in the absence of concanavalin A. Electron spin resonance studies have shown that the large lectin-glycoprotein complex (96,000 daltons) resides in a nonaqueous environment within the reverse micelle, perhaps at the surfactant, water-pool interface; hence, extraction of the large complex is feasible. The facilitative extraction has been extended to selective transport of peroxidase from a mixture of peroxidase and alkaline phosphatase (a nonglycosylated protein). This results in an efficient separation strategy with a separation factor of 16.
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PMID:Purification of glycoproteins by selective transport using concanavalin-mediated reverse micellar extraction. 137 45

The primary objective of the present study was to develop a method for quantifying the smooth muscle content of the prostate adenoma. A double immunoenzymatic staining technique was coupled with color assisted image analysis to determine the area density of the smooth muscle within the prostate adenoma. Eight males with symptomatic BPH underwent transrectal biopsy of the prostate. Four micron thick tissue sections were used for the double immunoenzymatic staining process. Rabbit anti-desmin and mouse anti-human prostatic acid phosphatase antibodies were used to selectively bind smooth muscle and prostatic epithelium, respectively. The two different tissue antigens were identified with peroxidase-antiperoxidase (PAP) and alkaline phosphatase-antialkaline phosphatase techniques. The alkaline phosphatase activity and peroxidase activity were developed with fast red and DAB chromogens. The BQ MEG IV Vista color system image analysis was used to discriminate color differences from the stained tissue sections. The thresholds were set to identify smooth muscle (dark brown), epithelium (red), fibrous tissue (pale brown), and glandular lumina (colorless). The mean area density of smooth muscle, fibrous tissue, glandular epithelium, and glandular lumina was 22%, 54%, 16%, and 9%, respectively. The present study suggests that a significant component of the prostate adenoma is smooth muscle. The application of this technique will be utilized to provide further insights into the role of smooth muscle in the pathogenesis and therapy of BPH.
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PMID:Quantifying the smooth muscle content of the prostate using double-immunoenzymatic staining and color assisted image analysis. 137 63

The purpose of this study was to assess the relationship of neuropeptide nerves and inflammatory leukocytes in PVG rats with adjuvant-induced arthritis. Substance P- and calcitonin gene-related peptide (CGRP)-immunoreactive nerves and inflammatory leukocytes were studied, using peroxidase (ABC) and/or alkaline phosphatase (APAAP) staining. Inflamed synovial tissue proper was infiltrated with neutrophils, ED1 macrophages and focal accumulations of CD2 T lymphocytes. In such tissue, the relationship between peptide-immunoreactive nerves and inflammatory cells was such that substance P and CGRP nerves were absent in heavily infiltrated villous synovial tissue, whereas healthy synovial tissue and non-inflammatory areas in adjuvant arthritic rats were innervated by substance P and CGRP nerves close to normal synovial tissue resident cells. In order to elucidate an eventual mechanism for lost immunoreactivity, healthy synovial tissue was exposed to chymotrypsin or oxygen derived free radicals (ODFR) in vitro. The former treatment caused total loss of immunoreactivity. These findings suggest that neuropeptides and neuropeptide containing nerves may be destroyed by locally produced proteolytic enzymes and various reactive oxygen species in the vicinity of inflammatory cells.
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PMID:Relationship between neuropeptide immunoreactive nerves and inflammatory cells in adjuvant arthritic rats. 137 4

Anti-desmin and anti-actin are commercially available antibodies that bind to smooth muscle. The present study was designed to compare the staining properties of anti-desmin and anti-actin in the human prostate in order to determine the optimal antibody for quantifying the smooth muscle content of the human prostate. Nineteen male subjects with symptomatic BPH underwent needle biopsy of the prostate. Double-immunoenzymatic staining was performed with peroxidase-anti-peroxidase (PAP) and alkaline phosphatase-anti-alkaline phosphatase (APAAP) techniques. Rabbit anti-desmin:mouse anti-human prostatic acid phosphatase and mouse anti-actin:rabbit anti-human prostatic acid phosphatase were utilized. Computer assisted color image analysis was performed using the Bioquant image analysis system. The percent area density of stroma and epithelium was independent of the antibodies used. The percent area density of smooth muscle in the anti-actin stained tissue sections was twofold greater than the anti-desmin stained tissue sections. A direct relationship was observed for the area density of smooth muscle (r = 0.71; P = 0.0006) and the area density of connective tissue (r = 0.82; P less than 0.001) determined from anti-desmin and anti-actin stained tissue sections. Anti-actin represents the optimal antibody for quantifying the area density of prostate smooth muscle. The reproducibility of the immunoenzymatic staining technique is inferred from the direct relationship observed for area density of epithelium between the different staining techniques.
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PMID:Anti-desmin vs. anti-actin for quantifying the area density of prostate smooth muscle. 137 9

A new simultaneous double immunostaining method has been optimized to localize the DNA synthesis marker bromodeoxyuridine (BrdU) and calcitonin gene-related peptide (CGRP) in endocrine cells of Bouin's-fixed, paraffin-embedded rat lung. Nuclease pre-treatment before immunostaining is compatible with optimal tissue morphology and CGRP antigenicity preservation. Nickel-enhanced development of avidin-biotin-peroxidase staining is used to show CGRP immunoreactivity in black and alkaline phosphatase-anti-alkaline phosphatase is applied to demonstrate incorporated BrdU in red. The present methodology could be useful for studies requiring detection of incorporated BrdU in cells producing regulatory peptides or other labile antigens.
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PMID:Simultaneous immunostaining method for localization of bromodeoxyuridine and calcitonin gene-related peptide. 137 32

Five murine monoclonal antibodies raised against Onchocerca volvulus cuticular extracts and termed MOVs (1-4 and 6) were selected based on reactivity with O. volvulus cryosections, and non-reactivity with cryosections of human skin and/or nodular tissue. Two others MOVs 5 and 7 reacted with both. Using the peroxidase-anti- peroxidase (PAP) histochemical method, the target epitopes of MOV 1 were located in the cuticle's basal and cortical layers, those of MOV 2 in the cortical layer; whilst MOV 3-7 stained the basal layer. A sandwich ELISA was then developed. The trapping polyclonal antibody was raised in rabbits utilising the same antigens as for preparation of the MOVs. Once captured on microtiter plates, target antigens were identified by the sequential binding of a MOV, followed by a goat anti-mouse globulin/peroxidase or alkaline phosphatase conjugate that catalysed a colorimetric reaction in the presence of appropriate substrates. In this system, MOV 1 emerged as the most specific and potent reagent capable of recognizing antigens of Onchocerca sp. with a minimal detection limit of 78 ng per test. MOV 1, failed to react with extracts of Loa Loa, Ascaris lumbricoides and Ascaris suum in the test. The developed assay relied on the use of MOV 1, required only 1 ml of urine or 0.05 ml serum. About 97.8% of the 47 urines and 50% of the 20 sera from patients studied gave positive results. Only 1 (3%) of 32 control urines and up to 80% of the 10 control sera studied tested positive, suggesting urine as a better specimen source.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:A monoclonal antibody-based immunodiagnostic assay for onchocerciasis. 138 17


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