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Biomedical subjects

H Harris

Publications and source records attributed to H Harris.

At least 127 records · Page 7Linked to original sources
PubMed1984

Studies of the biochemical and immunological properties of human neutrophil alkaline phosphatase with comparison to the established alkaline phosphatase isoenzymes.

A range of affinity column chromatographic procedures and various inhibitors have been used to compare human neutrophil alkaline phosphatase with the three established isoenzymes. The column chromatography studies have clearly distinguished neutrophil alkaline phosphatase from the intestinal isoenzyme. Inhibition studies with L-phenylalanine, L-homoarginine and levamisole have revealed a distinct pattern of inhibition for liver, kidney and neutrophil alkaline phosphatase which is quite different from the pattern shown by placental and intestinal alkaline phosphatase. Immunospecificity experiments with a monoclonal antibody raised to human liver alkaline phosphatase have shown that it cross reacts with alkaline phosphatase from kidney, bone and neutrophil. In all studies, neutrophil alkaline phosphatase has virtually identical properties to that of liver, kidney and bone alkaline phosphatase. This is strong evidence that neutrophil alkaline phosphatase is a product of the same structural gene which codes for the liver/bone/kidney group of human alkaline phosphatases.

Alkaline Phosphatase↗
PubMed1984

Monoclonal antibodies against placental-like and intestinal-like alkaline phosphatases in a malignant human cell line.

Monoclonal antibodies were raised against alkaline phosphatase (ALP) from the malignant human cell line Hep2, a derivative of HeLa, which was established from a cervical carcinoma. Two forms of ALP are found in Hep2 cells resembling, but not identical to, the normal placental and intestinal ALPs. Seven monoclonal antibodies were raised against Hep2 ALP. All of these reacted nearly identically with normal placental ALP and the placental-like ALP from Hep2 cells. Three of these antibodies exhibited reduced reactivities for either the common type 2 or 3 allelic variants of placental ALP. From these binding studies, the placental-like ALP from Hep2 cells appears to be a slightly modified form of the normal type 1 placental ALP. One of the seven antibodies bound to the adult and fetal intestinal ALPs, but more strongly to the adult form, suggesting that the Hep2 intestinal-like ALP is a form of the normal adult intestinal ALP.

Alkaline Phosphatase↗
PubMed1984

Purification of human adult and foetal intestinal alkaline phosphatases by monoclonal antibody immunoaffinity chromatography.

We have used the technique of monoclonal antibody immunoaffinity chromatography to purify adult and foetal intestinal alkaline phosphatases. Pure adult intestinal enzyme was obtained from a crude tissue extract with a single immunoaffinity chromatographic step in yields exceeding 95%. An additional ion-exchange chromatographic step was necessary for purification of the foetal enzyme, but yields still exceeded 70%. Experiments to optimize the efficiency of the monoclonal antibody immunoaffinity chromatography procedure suggest that the relative strength of binding of an antibody to its antigen is the most important factor to consider when constructing such columns. A column made from an antibody of too low an avidity will not retain the enzyme, while one of too high an avidity will make elution of enzyme in the active state difficult. A scheme is suggested for the application of this technique to a general approach to enzyme purification.

Adult↗
PubMed1984

Isoelectric-focusing of galactokinase in lens and other tissues.

Thin-layer isoelectric focusing was used to investigate galactokinase in lenses from humans, cows, rats, rabbits and also in other tissues (red cells, liver, kidney, brain, placenta) from these species. In each case the enzyme activity was present as a single isozyme. The isoelectric point was the same in lens, red cell and other tissues of the same species, but differed from species to species. Post-translational modification due to deamidation was not detected in the lens or red cell from these species. Galactokinase activities in whole tissue extract were determined in different tissues of the various species. There were wide variations in activity. In the adult human lens, both normal and cataractous, it was low. The findings indicate that isoelectric focusing of RBC galactokinase combined with specific enzyme staining allows one to monitor lens galactokinase in patients with cataracts.

Adult↗
PubMed1984

Structural analysis of human adult and fetal alkaline phosphatases by cyanogen bromide peptide mapping.

The adult and fetal forms of human intestinal alkaline phosphatase (ALPase; orthophosphoric-monoester phosphohydrolase, EC 3.1.3.1) are indistinguishable by a variety of analytical procedures. However, they differ electrophoretically and can be differentiated by binding studies with monoclonal antibodies. In this report, these two enzymes along with placental and liver ALPases are compared by the technique of CNBr peptide mapping, and the role of carbohydrate in generating these patterns is investigated. NaDodSO4/PAGE of CNBr digests of radiolabeled ALPases from fetal and adult intestine shows that these two isozymes share five of seven common-sized CNBr fragments. Placental ALPase shares only one common-sized fragment with either intestinal enzyme. Liver ALPase has no CNBr fragments in common with any of the others. These data indicate that fetal intestinal ALPase is not a heterodimer of one subunit each of intestinal ALPase and placental ALPase as has been postulated. CNBr digests of neuraminidase-treated enzymes reveal a change of mobility of only one CNBr band in each of fetal intestinal, placental, and liver ALPases, indicating the presence of sialic acid residues in these fragments. Periodic acid/Schiff reagent staining (specific for carbohydrate) of CNBr digests of fetal and adult intestinal ALPases reacts with only one band in each enzyme, which is the same band from the fetal enzyme shown to contain sialic acid. However, fetal and adult intestinal ALPases each contain at least one CNBr fragment of unique size that is apparently nonglycosylated.

Alkaline Phosphatase↗
PubMed1984

Two monoclonal antibodies recognizing determinants on human embryonal carcinoma cells react specifically with the liver isozyme of human alkaline phosphatase.

From a series of hybridomas that produced monoclonal antibodies reactive with the surface of human embryonal carcinoma cells, two that specifically recognized determinants of the liver/bone/kidney isozyme of alkaline phosphatase were isolated. They did not cross-react with the intestinal or placental isozymes. Phylogenetic studies revealed that both antibodies cross-reacted strongly with liver alkaline phosphatase from higher primates, but exhibited marked differences in their respective cross-reactions with liver alkaline phosphatase from other mammalian species.

Alkaline Phosphatase↗
PubMed1984

Identity of the neoplastic alkaline phosphatase as revealed with monoclonal antibodies to the placental form of the enzyme.

A panel of six monoclonal antibodies and a conventional polyclonal antibody raised against human placental alkaline phosphatase (ALP) were used to characterize the alkaline phosphatase detected by means of histochemistry on tumors of breast, ovary, lung, gastrointestinal tract, and kidney. Complete antigenic identity between the tumor ALP and the placental ALP was found only in one lung tumor. However, ten tumors reacted with the polyclonal antibody and some monoclonal antibodies, thus exhibiting partial identity with the placental ALP.

Adenocarcinoma↗
PubMed1984

Cytogenetic studies on recipients of allogeneic bone marrow using the sex chromosomes as markers of cellular origin.

In 45 patients whose donor was of unlike sex, the sex chromosomes were used as markers of the cellular origin of myeloid and lymphoid tissues after allogeneic bone marrow transplantation (BMT). Successful engraftment was characterized by the appearance of dividing donor cells in marrow within 2 weeks of grafting and in mitogen stimulated blood cultures by 3 weeks. Leukaemic relapse was identified in eight cases and was associated with different patterns of cellular origin of the myeloid and lymphoid tissues. At the time of relapse the marrow contained either a mixed population of normal donor and leukaemic recipient cells, or only recipient cells. Thus, in this series, leukaemic relapse was not found occurring in donor cells. The importance of defining the origin of cells in interphase as well as in metaphase was demonstrated. In all but one case, the dividing lymphoid population remained of donor origin during relapse.

Adolescent↗
PubMed1984

Alterations induced by glucose deprivation and tunicamycin in the kinetic parameters of hexose transport in hybrid cells.

Matched pairs of malignant and non-malignant hybrid cells were compared in their response to glucose deprivation and to tunicamycin. Glucose deprivation induced an increase in the maximum velocity in the malignant cells, but not in the non-malignant cells. The Michaelis constant of hexose uptake was largely unchanged by glucose deprivation except in the case of one melanoma derivative, PG19 G-, which showed a large increase in Michaelis constant when deprived of glucose. Tunicamycin increased the Michaelis constant of hexose uptake in both malignant and non-malignant cell lines. It is therefore possible that the Michaelis constant of hexose uptake is affected by the extent of glycosylation of one or more of the cell membrane glycoproteins.

Animals↗
PubMed1984

Soluble exopeptidases of bovine and human lens: characterization by electrophoresis.

Soluble exopeptidases present in bovine and human lenses were identified and characterized using starch gel electrophoresis separation followed by activity staining with an L-aminoacid oxidase/peroxidase system or a naphthylamine fluorescence system. Sixteen peptide and twelve naphthylamide substrates were used. The profile of substrate specificities for each electrophoretically separated exopeptidase was determined. Characterization also included the effects on activity of pH, EDTA, puromycin, and divalent cations. In addition, molecular weight determinations by gel filtration were made. Six bovine lens peptidases were identified including leucine aminopeptidase and dipeptidylpeptidase III and six human lens peptidase including dipeptidylpeptidase III. Strong homology in terms of substrate specificity and molecular weight was seen between bovine leucine aminopeptidase and one of the human peptidases previously designated peptidase "S". The findings indicate the diversity of exopeptidase available for polypeptide degradation in lens.

Animals↗
PubMed1984

Differentiation of human adult and fetal intestinal alkaline phosphatases with monoclonal antibodies.

Two forms of intestinal alkaline phosphatase have been recognized in humans. They are very similar in a number of biochemical and immunologic characteristics, but the exact genetic relationship between them remains unclear. To further study this problem, six monoclonal antibodies and a polyclonal rabbit antiserum to human fetal intestinal alkaline phosphatase have been produced. All of the monoclonal antibodies and the rabbit antiserum crossreact with adult intestinal alkaline phosphatase and with the intestinal-like alkaline phosphatase found in D98/AH-2 human tissue-culture cells. Four of the monoclonal antibodies and the rabbit antiserum crossreact with placental alkaline phosphatase, while none of the antibodies or the antiserum recognize liver or kidney alkaline phosphatase. Four of the monoclonal antibodies can distinguish between adult and fetal intestinal alkaline phosphatase in electrophoretic titration-binding studies, with the relative binding of adult enzyme being significantly greater than that of the fetal enzyme in each case. One of these antibodies, which also reacts with placental alkaline phosphatase, can distinguish the type 3 allelic variant of the placental enzyme from types 1 and 2. This indicates that the antibody detects a structural difference in the protein moiety of one of the allelic forms of the enzyme. These data suggest that adult and fetal intestinal alkaline phosphatases represent structurally distinct proteins, either encoded for by different genes or produced by differential processing of a common precursor molecule determined by a single gene.

Adult↗
PubMed1984

Use of monoclonal antibodies to assign phenotypes to placental alkaline phosphatase from cultured human cell lines derived from tumors.

Monoclonal antibodies were used to type placental alkaline phosphatase (ALP) from cell lines established from malignant human tumors by incubating ALP extracts from the cells with antibodies of different allelic specificities and separating free from bound enzyme on polyacrylamide gel electrophoresis. The HeLa-derived cell lines (Hep 2 and WISH) have the type 1 ALP phenotype, while a non-HeLa cell line (HT-3) has the type 2 ALP phenotype. This approach should prove of value for the phenotyping of enzymes and proteins with poorly resolved or altered electrophoretic patterns.

Alkaline Phosphatase↗
PubMed1983

Characterization of normal and abnormal variants of galactose-1-phosphate uridylyltransferase (EC 2.7.7.12) by isoelectric focusing.

Isoelectric focusing (IEF) in polyacrylamide gels has been used to study the isozymes of human galactose-1-phosphate uridylyltransferase (GALT) in erythrocytes and fibroblasts. In addition to the usefulness of IEF in differentiating normal, Duarte variant, and galactosemic homozygotes and heterozygotes, the ability of IEF to distinguish the residual GALT activity in two different galactosemic fibroblast lines and in revertants from them is demonstrated.

Cells, Cultured↗
PubMed1983

Tissue distribution of mammalian aldose reductase and related enzymes.

Activities of aldose reductase (AR) and related NADPH-dependent enzymes were examined in extracts of human, cat, dog, guinea pig, mouse, monkey, pig, rabbit, rat and sheep lenses and a variety of other tissues. The activity of the tissues against DL-glyceraldehyde, D-glucuronic acid, and 3-pyridinecarboxaldehyde (PCA) was determined. High glyceraldehyde:glucuronic acid activity ratios, a characteristic of aldose reductase, were found in all lenses, except from mouse. An analytical thin-layer isoelectric focusing system which separates the mammalian NADPH-dependent enzymes was developed. AR appears to be present as two or more isozymes in all mammalian lenses studied with the exception of mouse. Other tissues contain one or more isozymes which have the same isoelectric point and substrate specificity as the AR present in the lens of that species. This AR activity, however, may represent only a small proportion of the total NADPH reducing activity present. AR and HDH isozymes reduce the aromatic substrate, PCA, and thus have the general characteristics of an aldehyde reductase.

Alcohol Oxidoreductases↗