Purification of prostacyclin synthase from bovine aorta by immunoaffinity chromatography. Evidence that the enzyme is a hemoprotein.

SummaryTo reduce the risk of pathogenic virus transmission associated with the therapeutic administration of plasma-derived antihemophilic factor (FVIIIc), a process utilizing anti-FVIIIc immunoaffinity chromatography to isolate FVIIIc has been developed. In addition, the starting cryoprecipitate solution has been treated with an organic solvent/detergent mixture to inactivate lipid-enveloped viruses. A final ion exchange chromatography step is used to further remove contaminants, e.g., anti-FVIIIc antibody, potentially leached with FVIIIc during the immunoaffinity step. The purified FVTII is stabilized for lyophili-zation and storage by the addition of human albumin. The monoclonal anti-FVIIIc antibody used in the immunoaffinity step of the process is not detectable in the final preparation. Viral reduction studies performed at specific steps of the process demonstrate that 11 logs of human immunodeficiency virus (HIV) and greater than 4-5 logs of other lipid-enveloped viruses are inactivated within the first 30 s of exposure to the solvent/ detergent mixture and 4-5 logs of various model viruses, e. g. Endomyocarditis virus (EMC), are physically removed during washing of the immunoaffinity column. The lyophilized product is reconstituted using sterile water in a matter of seconds.The pharmacokinetics of Hemofil® M were compared to those obtained using a standard heat-treated concentrate (Hemofil® CT) in five severe factor VIII deficient hemophiliacs in a randomized, cross-over study. No statistically significant differences were observed in mean half life (p >0.6) or median recovery (p = 0.4) between the two preparations. No clinically significant adverse effects were observed in patients receiving either FVIII preparation.In addition, 43 patients at 18 different centers underwent pharmacokinetic studies, with a nominal dose of 50 u/kg FVIIIc Hemofil® M. The mean recovery was 103.6%, and the t 1/2 was 14.6 h. The recovery of FVIII in this group was as expected, providing an increase of assayed FVIII of approximately 2% per unit of FVTII/kg infused.Clinical trials using Hemofil® M have been initiated in 124 hemophilia A patients. The safety and efficacy of Hemofil® M has been established. To date, 0 of 60 patients tested have seroconverted to HIV. None of the previously untreated patients show clinical or laboratory evidence of Non-A, Non-B hepatitis (NANB), with 21 patients remaining negative as far as presence of antibodies to the Hepatitis C virus (a-HCV negative) at least 6 months after the initial infusion. There is no evidence of neoantigenicity, evidenced by seroconversion to murine antibody. An 8.7% (2 of 23) prevalence of anti-FVIIIc inhibitor development has been observed in previously untreated patients with FVIIIc⩽3%, receiving only the monoclonally purified solvent/ detergent treated FVIII concentrate while on study and on poststudy surveillance. All patients demonstrated clinical hemostasis following product use for either on demand bleeding or surgical prophylaxis.

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Milk and milk powder. Determination of aflatoxin M1 content. Clean-up by immunoaffinity chromatography and determination by thin-layer chromatography

10.3403/30043089 ◽

2005 ◽

Keyword(s):

Thin Layer ◽

Thin Layer Chromatography ◽

Milk Powder ◽

Immunoaffinity Chromatography ◽

Layer Chromatography

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Nitric Oxide in Life and Death of Neutrophils

Current Medicinal Chemistry ◽

10.2174/0929867326666181213093152 ◽

2019 ◽

Vol 26 (31) ◽

pp. 5764-5780 ◽

Cited By ~ 1

Author(s):

Svetlana I. Galkina ◽

Ekaterina A. Golenkina ◽

Galina M. Viryasova ◽

Yulia M. Romanova ◽

Galina F. Sud’ina

Keyword(s):

Nitric Oxide ◽

Cell Damage ◽

Protective Role ◽

Neutrophil Apoptosis ◽

High Biological Activity ◽

Life And Death ◽

Signalling Molecule ◽

Activated Neutrophils ◽

Cell Death Mechanisms ◽

Prostacyclin Synthase

Background: Nitric Oxide (NO) is a key signalling molecule that has an important role in inflammation. It can be secreted by endothelial cells, neutrophils, and other cells, and once in circulation, NO plays important roles in regulating various neutrophil cellular activities and fate. Objective: To describe neutrophil cellular responses influenced by NO and its concomitant compound peroxynitrite and signalling mechanisms for neutrophil apoptosis. Methods: Literature was reviewed to assess the effects of NO on neutrophils. Results: NO plays an important role in various neutrophil cellular activities and interaction with other cells. The characteristic cellular activities of neutrophils are adhesion and phagocytosis. NO plays a protective role in neutrophil-endothelial interaction by preventing neutrophil adhesion and endothelial cell damage by activated neutrophils. NO suppresses neutrophil phagocytic activity but stimulates longdistance contact interactions through tubulovesicular extensions or cytonemes. Neutrophils are the main source of superoxide, but NO flow results in the formation of peroxynitrite, a compound with high biological activity. Peroxynitrite is involved in the regulation of eicosanoid biosynthesis and inhibits endothelial prostacyclin synthase. NO and peroxynitrite modulate cellular 5-lipoxygenase activity and leukotriene synthesis. Long-term exposure of neutrophils to NO results in the activation of cell death mechanisms and neutrophil apoptosis. Conclusion: Nitric oxide and the NO/superoxide interplay fine-tune mechanisms regulating life and death in neutrophils.

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Prostacyclin Synthase Gene: Implication and Prevention of Cardiovascular Disease

Vascular Disease Prevention ◽

10.2174/1567270043404962 ◽

2004 ◽

Vol 1 (3) ◽

pp. 255-261

Author(s):

Tomohiro Nakayama

Keyword(s):

Cardiovascular Disease ◽

Prostacyclin Synthase ◽

Prevention Of Cardiovascular Disease

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Isolation and molecular cloning of prostacyclin synthase from bovine endothelial cells.

Journal of Biological Chemistry ◽

10.1016/s0021-9258(17)32104-x ◽

1994 ◽

Vol 269 (31) ◽

pp. 19897-19903

Author(s):

S. Hara ◽

A. Miyata ◽

C. Yokoyama ◽

H. Inoue ◽

R. Brugger ◽

...

Keyword(s):

Endothelial Cells ◽

Molecular Cloning ◽

Bovine Endothelial Cells ◽

Prostacyclin Synthase

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Rapid and simultaneous purification of aflatoxin B1, zearalenone and deoxynivalenol using their monoclonal antibodies and magnetic nanoparticles

Toxicological Research ◽

10.1007/s43188-020-00083-w ◽

2021 ◽

Author(s):

Hyuk-Mi Lee ◽

Hwan-Goo Kang

Keyword(s):

Monoclonal Antibodies ◽

Magnetic Nanoparticles ◽

Aflatoxin B1 ◽

Animal Feed ◽

Immunoaffinity Chromatography ◽

The Novel ◽

Buffer Solution ◽

Purification Method ◽

Recovery Rates ◽

Single Spike

AbstractTo develop a new simple and simultaneous purification method for mycotoxins in feeds and grains, magnetic nanoparticles (MNPs) conjugated with monoclonal antibodies (mAbs) against mycotoxins were used to separate aflatoxin B1 (AFB1), zearalenone (ZEA) and deoxynivalenol (DON). For a single spike of each mycotoxin into the buffer solution (16% MeOH in PBS), mean recoveries were 93.1–95.0% for AFB1 (5–20 ng/mL spiked), 87.2–96.0% for ZEA (125–500 ng/mL spiked) and 75.2–96.9% for DON (250–1,000 ng/mL spiked) by HPLC and ELISA. Recoveries of AFB1 (20 ng/mL) and ZEA (500 ng/mL) simultaneously spiked into the buffer solution were 87.0 and 99.8%, respectively. Recovery rates of AFB1/DON and DON/ZEA spiked simultaneously were 86.2%/76.6% and 92.0%/86.7%, respectively, at concentrations of 20 ng/mL AFB1, 500 ng/mL ZEA, and 1,000 ng/mL DON. Recoveries using the novel mAb–MNP conjugated system in a buffer solution simultaneously spiked with AFB1, ZEA and DON were 82.5, 94.6 and 73.4%, respectively. Recoveries of DON in animal feed were 107.7–132.5% at concentrations of 250–1,000 ng/g spiked in feed. The immunoaffinity chromatography (IAC) clean-up method was compared with the purification method using novel mAb–MNP. After fortification of animal feed with AFB1 (5, 10 and 20 ng/g feed) and ZEA (125, 250 and 500 ng/g feed), AFB1 and ZEA were purified using both the methods. In the case of the novel mAb-MNP conjugated system, mean recoveries for AFB1 were 89.4, 73.1 and 88.3% at concentrations of 5, 10 and 20 ng/g feed, respectively. For ZEA, mean recoveries were 86.7, 85.9 and 79.1% at concentrations of 125, 250 and 500 ng/g, respectively. For IAC purification, recoveries were 42.9–45.1% for AFB1 and 96.8–103.2% for ZEA. In conclusion, the present purification method using monoclonal antibodies conjugated to MNPs can be used for simple and simultaneous purification of mycotoxins from feed and maize.

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