Methods for the Determination of the Interferon-Induced Enzyme 2'-5' Oligoadenylate Synthetase in Mononuclear Blood Cells

Bruchelt, Gernot; Beck, James H.; Schilbach-Stückle, Karin; Kościelniak, Ewa; Treuner, J.; Niethammer, D.

doi:10.1515/cclm.1987.25.12.879

Cited by 3 publications

(6 citation statements)

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“…Mononuclear blood cells (MNBC) from healthy persons were isolated from heparinized blood by centrifugation using Lymphoprep (Nyegaard, Oslo, Norway) as described [29]. The neuroblastoma cell line SK-N-SH was obtained from the American Type Culture Collection (ATCC).…”

Section: Cells and Preparation Of Cell Lysatesmentioning

confidence: 99%

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Determination of 2—5A synthetase and 2—5A phosphodiesterase in neuroblastoma cells by analytical capillary isotachophoresis: Effects of cytokines and comparison with radioenzymatic methods

et al. 1994

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2-5A Synthetase and 2-5A phosphodiesterase were determined by analytical capillary isotachophoresis in comparison to radioenzymatic methods. By means of isotachophoretic analysis, a frequently used radioenzymatic 2-5A synthetase assay was optimized and the results of both assays were compared. Using the isotachophoretic assay the influence of interferon-related cytokines (tumor necrosis factor-alpha and interleukin-2) on 2-5A synthetase induction in neuroblastoma cells was estimated. In contrast to mononuclear blood cells, the tumor necrosis factor induced 2-5A synthetase in these cells. 2-5A Phosphodiesterase was determined using an isotachophoretic assay and a radioenzymatic method. Degradation of A2'p5'A2'p5'A (trimeric form of 2-5A core) was measured by isotachophoresis whereas degradation of a mixture of phosphorus-32 labeled 2-5A cores was registered by radioenzymatic assay. Activity of 2-5A phosphodiesterase was only insignificantly enhanced by interferon in mononuclear blood and neuroblastoma cells. In contrast to the radioenzymatic assays, an accurate determination of 2-5A synthetase as well as of 2-5A phosphodiesterase is possible using the isotachophoretic method because the reactions are followed by measuring the substrates ATP and A2'p5'A2'p5'A, respectively.

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Section: Cells and Preparation Of Cell Lysatesmentioning

confidence: 99%

“…Determination of 2-5A synthetase and 2-5A PDE by radioenzymatic assays 2-5A Synthetase was determined using a slightly modified assay according to Schattner [27], as described in detail in [29]. A simplified flow diagram of this assay is given in Fig.…”

Section: 3mentioning

confidence: 99%

Determination of 2—5A synthetase and 2—5A phosphodiesterase in neuroblastoma cells by analytical capillary isotachophoresis: Effects of cytokines and comparison with radioenzymatic methods

et al. 1994

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“…Leukocytes were isolated by centrifugation using Lymphoprep (Nyegaard, Oslo, Norway) as described (23 The radioimmunoassay of EIKEN (Immunological Laboratory Kita-ku, Tokyo, Japan), provided by Mecconti, Bad Homburg, Germany, was used. The test was carried out according to the manufacturer's instructions:…”

Section: Blood Samplesmentioning

confidence: 99%

“…This enzyme was determined by a slightly modified procedure of the assay according to Schattner (25), and is described in detail elsewhere (23).…”

Section: Determination Of 2'-5'-oligoadenylate Synthetase In Leukocytesmentioning

confidence: 99%

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Determination of 2´-5´-Oligoadenylate Synthetase in Serum and Peripheral Blood Mononuclear Cells before and after Subcutaneous Application of Recombinant Interferon beta and gamma

Bruchelt¹,

Fierlbeck²,

Schiebel³

et al. 1992

Clinical Chemistry and Laboratory Medicine

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Establishment of a nonradioactive assay for2’-5’ oligoadenylate synthetase and its application in chronic hepatitis C patients receiving interferon-α

Tong¹

1998

WJG

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AIM:To establish a nonradioactive assay for 2'-5' oligoadenylate synthetase (2-5 AS) and to measure the 2-5AS in peripheral blood mononuclear cell (PBMC) extracts of patients with chronic hepatitis C before IFN-alphainjection, 24 hours and one month after the first injection.METHODS: 2-5AS in cell extracts of PBMCs from 10 normal persons and 15 chronic hepatitis C patients were determined with PEI cellulose thin-layer chromatography.RESULTS: The assay of 2-5AS in human PBMC was found to be rapid, sensitive, specific and reliable. The 2-5AS activity of PBMC in normal persons was in a quite low level (2.0%), and it was increased about ten-folds after stimulation of IFN (19.7%), (P < 0.01). In 15 chronic hepatitic C patients, the basal levels of 2-5AS before IFN treatment were higher than those of normal persons, being much higher in the group showing poor response to IFN treatment, but 24h after the first injection of IFN-alphathe 2-5AS level showed a more rapid and much greater rise in those patients with a good response.CONCLUSION: 2-5AS may be a useful parameter of biological response during the IFN therapy.

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Methods for the Determination of the Interferon-Induced Enzyme 2'-5' Oligoadenylate Synthetase in Mononuclear Blood Cells

Cited by 3 publications

References 10 publications

Determination of 2—5A synthetase and 2—5A phosphodiesterase in neuroblastoma cells by analytical capillary isotachophoresis: Effects of cytokines and comparison with radioenzymatic methods

Determination of 2—5A synthetase and 2—5A phosphodiesterase in neuroblastoma cells by analytical capillary isotachophoresis: Effects of cytokines and comparison with radioenzymatic methods

Determination of 2´-5´-Oligoadenylate Synthetase in Serum and Peripheral Blood Mononuclear Cells before and after Subcutaneous Application of Recombinant Interferon beta and gamma

Establishment of a nonradioactive assay for2’-5’ oligoadenylate synthetase and its application in chronic hepatitis C patients receiving interferon-α

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