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HLA-DR and -DQ Serotyping
Published in M. Kam, Jeffrey L. Bidwell, Handbook of HLA TYPING TECHNIQUES, 2020
Supernatants normally should be stored neat and can be stored at 4°C with 0.1% sodium azide to avoid loss of activity. Locally, we have found that if a supernatant is relatively strong loss of activity is negligible following one cycle of freezing and thawing; repeated freezing and thawing should be avoided.
Decarboxylation of 7-−14 C-Benzoic Acid
Published in Robert A. Greenwald, CRC Handbook of Methods for Oxygen Radical Research, 2018
Gary W. Winston, Arthur I. Cederbaum
Detailed methods for the preparation of rat liver microsomes are described elsewhere in this volume. The standard reaction mixture consists of 100 mM potassium phosphate, pH 7.4, 10 mM MgCl2, 0.3 mM NADP+, 10 mM glucose 6-phosphate, 7 units of glucose 6-phosphate dehydrogenase, 0.1 mM EDTA, 1 mM sodium azide, about 5 mg of microsomal protein, and varying amounts (e.g., 5 to 30 mM of sodium benzoate, containing tracer amounts of 7-14C-benzoate (specific activity 5.5 μ Ci/mmol) in a final volume of 3 mℓ. The addition of EDTA increases the decarboxylation of benzoate, probably by chelating iron in the system. Azide is added to inhibit the activity of catalase, which is present as a contaminant in isolated microsomal preparations. The reaction is initiated by the addition of glucose 6-phosphate plus glucose 6-phosphate dehydrogenase and terminated by the addition of 0.3 mℓ of 70% perchloric acid. Under these conditions, a typical 20-min experiment yields counts which are at least 30 times greater than the background counts.
Ceruloplasmin
Published in René Lontie, Copper Proteins and Copper Enzymes, 1984
Manabe et al.275,276 observed a species with an absorption maximum at 420 nm, which occurs during the reoxidation of reduced Cp. The maximum intensity occurred after 200 msec, when about 70% of the 610-nm and practically all 330-nm absorption had returned. The decay rate of this species was decreased by azide to the same extent as the inhibition of the enzyme. In contrast, azide had no influence on the reduction of type-1 coppers or reappearance of the 610-nm absorption after reduction. The 420-nm intermediate might be the explanation for the overshoot in 340-nm absorption observed in the reoxidation experiments by Carrico et al.183 There are several indications that this species is an oxygen intermediate, e.g., the pseudo-first-order rate constant for the appearance of the species increases with oxygen concentrations.276 By rapid freezing Bränden and Deinum277 were able to study the transient EPR signal appearing during reoxidation. They found it to be similar to the one observed in fungal laccase, which was broadened by substituting 16O2 with 17O2. The line shape is extremely sensitive to pH changes between 4.4 and 6.6 indicating more than 1 proton equilibrium. The signal decayed in parallel as the type-2 Cu(II) reappeared. It should be added that the signal appears also in type-2 copper-depleted laccase which suggests that the type-3 coppers are responsible for oxygen binding, by analogy to the situation in tyrosinase (see Volume II, Chapter 7).
Enzyme/pH dual stimuli-responsive nanoplatform co-deliver disulfiram and doxorubicin for effective treatment of breast cancer lung metastasis
Published in Expert Opinion on Drug Delivery, 2023
Peifu Xiao, Xiaoguang Tao, Hanxun Wang, Hongbing Liu, Yupeng Feng, Yueqi Zhu, Zhengzhen Jiang, Tian Yin, Yu Zhang, Haibing He, Jingxin Gou, Xing Tang
Subsequently, the uptake mechanism of DSF-DOX NPs was investigated. As we know, nanoparticles could enter the tumor cells via endocytosis [58,59]. The endocytosis of nanoparticles into tumor cells is generally energy-dependent, this process can be divided into different types including caveolae-mediated endocytosis, clathrin-mediated endocytosis, and macropinocytosis [60,61]. Sodium azide is an electron transfer inhibitor that prevents energy production in cells. Chlorpromazine and philippine are inhibitors of clathrin-mediated endocytosis and caveolae-mediated endocytosis, respectively. Protamine can inhibit cell pinocytosis. As shown in Figure 6d, after pretreating 4T1 cells with sodium azide and chlorpromazine, compared with the control group, the drug fluorescence intensity in 4T1 cells decreased to 22.50% and 41.07%, respectively, indicating that the endocytosis of DSF-DOX NPs is an energy-dependent clathrin-mediated endocytosis. Meanwhile, the drug fluorescence intensity of the protamine group decreased to 67.85% compared with the control group, suggesting that cellular uptake of DSF-DOX NPs also involves the pinocytosis process. In contrast, the drug fluorescence intensity of the philippine group was still 90.01%. This result proved that caveolae-mediated endocytosis has little effect on the uptake process of DSF-DOX NPs. Taken together, clathrin-mediated endocytosis played the most vital role during the intracellular uptake process of DSF-DOX NPs.
The vitamin B12 analog cobinamide ameliorates azide toxicity in cells, Drosophila melanogaster, and mice
Published in Clinical Toxicology, 2023
John Tat, Stephen C. Chang, Cole D. Link, Suelen Razo-Lopez, Michael J. Ingerto, Behdod Katebian, Adriano Chan, Hema Kalyanaraman, Renate B. Pilz, Gerry R. Boss
The azide (N3-) anion is available in a variety of forms, most commonly as sodium azide (NaN3). Azide is used in several industries with over 1000 tons of NaN3 produced annually [1]. Azide is highly toxic, with the human lethal dose of sodium azide estimated to be ∼700 mg or ∼10 mg/kg [2]. Fortunately, azide poisoning is rare with 156 reported cases worldwide between the years 2000 and 2020 [3]. However, this infrequency may prove detrimental in a mass casualties event, such as an industrial incident or terrorist attack, as most medical personnel will not have encountered an azide-poisoned patient and therefore not be well-informed about azide toxicity and treatment options. A major terrorist attack is possible, since azide may be purchased through online retailers, and it has been used in several planned and executed attacks [4–9]. On several occasions, azide was used to poison communal beverages, leading to high casualties and highlighting the potential of azide as a terrorist weapon [10–14]. Moreover, azide is a common suicidal agent, especially among laboratory workers, likely due to its common presence in research laboratories [2,3,15].
Sodium azide poisoning: a narrative review
Published in Clinical Toxicology, 2021
John Tat, Karen Heskett, Shiho Satomi, Renate B. Pilz, Beatrice A. Golomb, Gerry R. Boss
Azide has several mechanisms of toxicity. At the cellular level, it inhibits mitochondrial cytochrome C oxidase and catalase [16,17]. The former enzyme is part of complex IV in the mitochondrial electron transport chain and the latter enzyme detoxifies hydrogen peroxide to water and oxygen. Thus, azide can reduce ATP synthesis and cause oxidative stress, the latter due both to mitochondrial electron leakage and reduced catabolism of reactive oxygen species. Cyanide also inhibits cytochrome C oxidase, and azide and cyanide are especially toxic to cells with high respiratory rates, such as neurons and cardiomyocytes. At the organismal level, azide is a potent vasodilator and inhibits platelet aggregation, likely via conversion to nitric oxide. Azide generates nitric oxide in vitro in erythrocytes, platelets, and isolated blood vessels, and recently, nitrosyl-hemoglobin was found in the blood of mice that had received azide [18]. Cytochrome C oxidase inhibition and nitric oxide generation likely underlie the hypotension, myocardial and respiratory failure, and metabolic acidosis that occur in azide poisoning.