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Biomedical subjects

M Totani

Publications and source records attributed to M Totani.

At least 19 recordsLinked to original sources
PubMed1998

Liver-type arginase in serum during and after liver transplantation: a novel index in monitoring conditions of the liver graft and its clinical significance.

We quantified liver-type arginase in sera of 47 patients undergoing partial liver transplantation with use of an ELISA method. The level of liver-type arginase fluctuated slightly beyond the normal range in successful liver recipients, while it changed more drastically or precipitously in unsuccessful ones, accompanying or unaccompanying elevation of AST and ALT levels. A higher elevation pattern of the arginase level (above 100 ng ml-1) was observed in each of the unsuccessful recipients with critical condition, except for one patient. Other hepatic markers (LDH, ALP, and T-BIL) remained relatively unchanged until the terminal stage of deceasing patients. The finding that the liver-type arginase emerged in large quantity in the blood stream immediately after reperfusion of the liver graft indicates that the enzyme leaks out of hepatocytes damaged, presumably, by storage in the absence of circulation. A half-life of the liver-type arginase in the human blood was estimated to be 1 h, that is clearly shorter than that of AST. The short half-life of the arginase appears to be ascribable, at least partly, to formation of an immune complex with circulating autoantibody which appears in many liver recipients. These results suggest that liver-type arginase behaves uniquely in the serum among many hepatic enzymes, and could serve as a distinct marker of hepatic lesions, particularly during and after liver transplantation.

Adolescent↗
PubMed1998

Sensitive enzyme immunoassay for anti-beta-lactoglobulin IgG in serum.

A sensitive enzyme immunoassay for anti-beta-lactoglobulin immunoglobulin G (IgG) in serum is described. Serum containing anti-beta-lactoglobulin IgG was reacted simultaneously with 2,4-dinitrophenyl-bovine serum albumin-beta-lactoglobulin conjugate and beta-lactoglobulin-peroxidase conjugate. The complex formed from the three components was trapped onto polystyrene balls coated with anti-2,4-dinitrophenyl group IgG, eluted with epsilon N-2,4-dinitrophenyl-L-lysine and transferred to polystyrene balls coated with anti-human IgG.gamma-chain IgG. Bound peroxidase activity was determined by fluorometry. This enzyme immunoassay was 100- to 1000-fold more sensitive and more reliable than the enzyme-linked immunosorbent assay (ELISA). Anti-beta-lactoglobulin IgG was detected in 91% of healthy subjects using this method.

Antibodies↗
PubMed1996

Structural organization and tissue-specific expression of the gene encoding rat cysteine dioxygenase.

Cysteine dioxygenase (CDO) is a key enzyme involved in the metabolism of L-cysteine. Genomic clones containing the 5'-flanking sequence of the rat CDO gene were isolated and characterized. The CDO gene spanned about 15 kb, and comprised 5 exons. All boundaries between the exons and introns matched the GT/AG rule. The major transcription start point (tsp) was A at 213 bp upstream from the ATG codon. The 5'-flanking region contained a TATA-box-like sequence and putative cis-acting regulatory elements. The 3' end of CDO was polyadenylated at several sites. Northern blots of RNA from rat tissues revealed the highest CDO mRNA level in the liver. Significant levels were observed in the kidney, lung and brain, implying tissue-specific differences in CDO promoter function.

Animals↗
PubMed1995

A chemiluminescence-flow injection analysis of serum 3-hydroxybutyrate using a bioreactor consisting of 3-hydroxybutyrate dehydrogenase and NADH oxidase.

We describe a simple method for the highly sensitive chemiluminescence--flow injection analysis of 3-hydroxybutyrate in serum using a bioreactor column consisting of the two immobilized enzymes, 3-hydroxybutyrate dehydrogenase and NADH oxidase. The method was based on measuring the level of chemiluminescence formed by the reaction of a luminol-hexacyanoferrate mixture with hydrogen peroxide. The hydrogen peroxide was produced by the NADH oxidase reaction from NADH which was formed in the conversion of 3-hydroxybutyrate to acetoacetate by the 3-hydroxybutyrate dehydrogenase reaction. Among three immobilized enzyme columns, a coimmobilized, small 3-hydroxybutyrate dehydrogenase/NADH oxidase bioreactor alone (2 x 20 mm i.d.) readily hydrolyzed all of the injected 3-hydroxybutyrate into acetoacetate, although 3-hydroxybutyrate dehydrogenase catalyzed the reversible reaction. The present method generated linearity of the data up to 1.5 mM 3-hydroxybutyrate with satisfactory precision, reproducibility, and accurate reaction recoveries. The results from 3-hydroxybutyrate correlated satisfactorily with those obtained by other well-established methods. The coimmobilized 3-hydroxybutyrate dehydrogenase/NADH oxidase reactor unit showed good operational stability over a 5-week period, during which it was repeatedly used for 1500 analyses.

3-Hydroxybutyric Acid↗
PubMed1995

[Lactic acid].

Explore the source record for details and available documents.

Acidosis, Lactic↗
PubMed1993

Enzyme immunoassay of liver-type arginase and its potential clinical application.

We developed an efficient enzyme-linked immunosorbent assay (ELISA) system for measurement of human liver-type arginase in serum. A conjugate of the Fab' fragment of anti-human liver (recombinant) arginase IgG and horseradish peroxidase was used as the second antibody. This assay is highly specific, sensitive, and reproducible, enabling us to detect arginase at concentrations as low as several micrograms per liter without any prior processing of serum. The reaction is linear up to 200 micrograms/L. The arginase concentration in serum, as determined by this method, increased markedly and temporarily at the time of surgical operation or later injury to the liver. The increase was accompanied or followed by increases in serum concentrations of aspartate aminotransferase, alanine aminotransferase, and lactate dehydrogenase, suggesting that the arginase emerged from damaged hepatocytes. In view of a limited tissue distribution of liver-type arginase, our ELISA system may be useful in diagnosis of various hepatic disorders as well as follow-up of postoperative conditions of patients.

Adolescent↗
PubMed1992

Substrate specificity and distribution of UDP-GalNAc:sialylparagloboside N-acetylgalactosaminyltransferase in the human stomach.

The detailed substrate specificity of the UDP-GalNAc:sialylparagloboside N-acetylgalactosaminyltransferase to form the Sd(a+) blood group active carbohydrate determinant GalNAc beta 1-4(NeuAc alpha 2-3)Gal was studied using a membrane fraction prepared from human gastric fundic mucosa. Various sialosylated oligosaccharides and gangliosides were examined as acceptor substrates. Oligosaccharide substrates were fluorescence-labelled with 2-aminopyridine, and the transferase activity was quantified by h.p.l.c. using a reversed-phase column. The structures of the products were determined by glycosidase degradation and proton n.m.r. 3'-Sialyl-lactose (II3NeuAcLac), 3'-sialyl-lactotetraose (IV3NeuAcLc4), and 3'-sialyl-lactoneotetraose (IV3NeuAcnLc4) were good substrates for the beta 1-4GalNAc transferase in gastric fundic mucosa, but 6'-sialyl-lactoneotetraose (IV6NeuAcnLc4) or 6'-sialyl-lactose (II6NeuAcLac) were not. Gangliosides with a terminal NeuAc alpha 2-3Gal residue such as GM3, sialylparagloboside, GM1b and GD1a were also studied. The activity of beta 1-4GalNAc transfer to sialylparagloboside was much higher than that to GM2, GM1b or GD1a in spite of them having the same terminal residue. Measurement of the activity of the beta 1-4GalNAc transferase in biopsy specimens demonstrated that the activity was localized in gastric fundic mucosa and was absent in pyloric mucosa, intestinal metaplasia and gastric cancer tissue. Thus the beta 1-4GalNAc transferase present specifically in fundic mucosa required a NeuAc alpha 2-3Gal residue connected to either type-1-chain or type-2-chain oligosaccharides. In glycolipids, the acceptor specificity was restricted to NeuAc alpha 2-3Gal beta 1-4GlcNAc because the NeuAc alpha 2-3Gal beta 1-3GalNAc structure in ganglio-series glycolipids was not a good acceptor substrate.

Aminopyridines↗
PubMed1992

Use of various types of column reactors for flow-injection analysis.

Two or three different kinds of immobilized enzymes can be aligned in a minireactor so that sequential enzymatic reactions are carried out from upstream to downstream during flow-injection analysis. A lactate oxidase-catalase reactor, used as precolumn for removing pre-existing lactate in serum before the lactose dehydrogenase (LDH) reactions, was useful for the determination of serum LDH activity, which did not require any blank correction. A sequential glutamate dehydrogenase-glutamate oxidase reactor was also useful for a novel chemiluminometric determination of ammonia. On the other hand, a co-immobilized creatininase-creatinase-sarcosine oxidase reactor, in spite of containing creatininase which catalyses the reversible reaction, was the most efficient for the determination of serum creatinine.

Ammonia↗
PubMed1992

[Biosensing techniques for the laboratory medicine].

Biosensing techniques are utilized in micro analysis of the biological constituents with an integrated device. Their applications to laboratory medicine are of great interest. Recently, several types of micro electrodes or ion sensitive field effect transistors (ISOFET) were developed. For fabrication of the biosensor selection of the proper bio-element and transducer to assay our sample, is important. In this article we will indicate some items on the future prospect of biosensing techniques for laboratory medicine.

Biosensing Techniques↗
PubMed1991

Determinations of lactate and lactate dehydrogenase activity in serum with the flow injection analysis system involving immobilized enzyme column and chemiluminescence.

The methods for the highly sensitive flow injection analysis of lactate and lactate dehydrogenase (LDH) activity in serum using immobilized enzymes in column form and chemiluminescence detection which does not require a blank correction are described. The methods were based on the determination of chemiluminescence formed by the reaction of a luminol-ferricyanide mixture with hydrogen peroxide. This hydrogen peroxide was produced by the lactate oxidase (LOD) reaction from lactate, which was in serum or was produced by the action of LDH in serum. The action of LDH in a flow injection analysis system was performed for 2 min in an incubation coil placed parallel to the substrate-buffer line between the LOD column and the LOD/catalase column. Endogenous lactate in serum was removed by an immobilized LOD/catalase column prior to the action of LDH. The present method gave perfect linearity of the data up to 5.6 mmol/liter for lactate and 1840 IU/liter for LDH activity with satisfactory precision, reproducibility, and accurate reaction recoveries. The results from the lactate and LDH activity correlated satisfactorily with those obtained by other well-established methods.

Autoanalysis↗
PubMed1991

Absence of erythrocyte arginase protein in Japanese patients with hyperargininemia.

In Japan, hyperargininemia has been reported in only 5 unrelated families and four patients are alive at present. In this study we examined arginase protein in erythrocytes of these Japanese patients using two analytical methods of immunoblotting and two-dimensional gel electrophoresis. Immunoblotting study with anti-E. coli-expressed human liver arginase rabbit IgG revealed lack of cross-reacting materials in the erythrocyte lysates from these patients. On two-dimensional gels, arginase protein was detected in any control subject, but it was completely absent in all the patients studied. These results suggest that either arginase protein in erythrocytes is not produced or it is structurally labile in these patients.

Adult↗
PubMed1990

Expression of human liver arginase in Escherichia coli. Purification and properties of the product.

Arginase is an enzyme that catalyses the hydrolysis of arginine to urea and ornithine. It is abundantly present in the liver of ureotelic animals (i.e. those whose excretion is characterized by the excretion of uric acid as the chief end-product of nitrogen metabolism), but its purification has hitherto not been simple, and the yield not high. Starting with a partially truncated cDNA for human liver arginase recently made available, we constructed an expression plasmid that had tandemly linked tac promotors placed upstream of a full-length cDNA. By selecting Escherichia coli strain KY1436 as the host micro-organism, we established an efficient system for the production of human liver arginase protein. Chromatographies on CM-Sephadex G-150, DEAE-cellulose and Sephadex G-150, followed by preparative agar-gel electrophoresis, yielded 10 mg of apparently homogeneous enzyme protein from 1 g (wet wt.) of E. coli cells. E. coli-expressed human liver arginase had chemical, immunological and most catalytic properties indistinguishable from those of purified human erythrocyte arginase. However, E. coli-expressed arginase was a monomer of Mr 35,000, whereas the purified erythrocyte arginase was trimer of Mr 105,000. They differed also in pH- and temperature-stabilities. Gel-filtration experiments with these two purified arginases under various conditions, as well as with unfractionated human liver and erythrocyte cytosol preparations, indicated that the native form of human arginase should be of Mr 35,000, and that the trimeric appearance of human erythrocyte arginase after purification was an artifact of the purification procedures. It was thus concluded that, in Nature, the liver and erythrocyte arginases are identical proteins.

Amino Acid Sequence↗
PubMed1990

A chemiluminometric method for NADPH and NADH using a two-enzyme bioreactor and its application to the determination of magnesium in serum.

Chemiluminometric methods are described for the automated flow injection analysis of NADPH and NADH using an immobilized enzyme column reactor and serum magnesium. This application is for the clinical analysis of NADPH and NADH. The reactor for NADPH and NADH contains immobilized L-glutamate dehydrogenase and L-glutamate oxidase, and that for serum magnesium immobilized hexokinase, glucose-6-phosphate dehydrogenase, L-glutamate dehydrogenase and L-glutamate oxidase. When the sample is introduced into the four-enzyme bioreactor, hydrogen peroxide is produced in proportion to the concentration of serum magnesium by the successive reactions. A co-immobilized hexokinase/glucose-6-phosphate dehydrogenase/glutamate dehydrogenase column reactor gave better efficiency compared with an enzyme column which was prepared by packing co-immobilized hexokinase/glucose-6-phosphate dehydrogenase and immobilized glutamate dehydrogenase to make two layers. Magnesium in serum was determined with 1 microL of the sample without carry-over and for an assay time of approximately 15 s. The present method is sensitive (detection limit 0.1 nmol) because Mg2+ is recycled in a column, and gives perfect linearity of the data up to 3.0 mmol/L with satisfactory precision, reproducibility, and accurate reaction recoveries.

Amino Acid Oxidoreductases↗
PubMed1990

A flow injection analysis system involving immobilized NADH oxidase in column form for clinical analysis.

A highly sensitive FIA system for chemiluminometric determination of reduced coenzyme, NADH, was developed, using immobilized NADH oxidase from Brevibacterium ammoniagenes. The enzyme catalyzed the oxidation of NADH generating hydrogen peroxide which emitted chemiluminescence when mixed with luminol and potassium ferricyanide. The immobilized enzyme reactor was a mini-column, measuring 1 or 2 mm in inner diameter and 20 mm in length, and the sample volume was only 1 microliter per assay, with a feeding speed of one sample per min and a lowest detection limit of 10 pmol NADH. A FIA system was also developed for the determination of magnesium in human serum, using an enzyme column reactor with simultaneously coimmobilized hexokinase, D-glucose-6-phosphate dehydrogenase, and NADH oxidase. The performance of the system was as satisfactory as a routine colorimetric assay, but with much higher sensitivity.

Colorimetry↗
PubMed1990

Monoclonal antibody inhibiting creatine kinase MM3 but not isoform MM1.

Monoclonal antibody CKM-G01 inhibited greater than 99% of the activity of porcine and human creatine kinase(CK)-MM isoenzyme purified from muscle. However, it inhibited only 54% of CK-MM in human serum. Chromatofocusing of serum CK-MM showed that CKM-G01 inhibited 100% of MM3 but not isoform MM1. CKM-G01 inhibited CK-MM2 by 57%. CKM-G01 specifically inhibited only the original CK-M subunit and not the subunit modified by removal of C-terminal lysine by carboxypeptidase N. CKM-G01 can be used for assay of CK isoforms. We devised a new diagnostic reagent involving it, which requires no analytical separation of isoforms, based on the immunoinhibition method, and applied it to early diagnosis of acute myocardial infarction. The "inhibition index," (inhibited CK activity/total CK activity) x 100, increased more rapidly than did total CK and CK-MB. Evidently this diagnostic reagent can be used for easy, early diagnosis of acute myocardial infarction.

Antibodies, Monoclonal↗