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Fluorescence in Histochemical Reactions
Published in Victoria Vladimirovna Roshchina, Fluorescence of Living Plant Cells for Phytomedicine Preparations, 2020
Victoria Vladimirovna Roshchina
Today, the main indirect way to determine acetylcholine in plant cells using a fluorescent method is to assay the presence of cholinesterase (Figure 4.1). One may determine the reactions with coumarinylphenylmaleimide (Parvari et al. 1983) or with the red analog of Ellman reagent (2,2-dithio-bis-(p-phenyleneazo)-bis-(1-oxy-8-chlorine-3,6)-disulfuric acid in the form of a sodium salt) for a fluorescent product with thiocholine (Roshchina et al. 1994; Roshchina 2001). The first candidate for histochemical application was the flavonoid derivative coumarinylphenylmaleimide (Parvari et al. 1983), which has a maximum of 390 nm in the excitation curve and 473 nm in the emission spectrum. After treatment with the second reagent, a blue compound was formed on the surface of pollen from various species and fluoresced with a maximum of 500–510 nm (Roshchina et al. 1994; Roshchina 2001), while untreated cells of Hippeastrum hybridum pollen, for example, usually emitted with a small maximum of 480 nm if at all.
Reactivities of Amino Acids and Proteins with Iodine
Published in Erwin Regoeczi, Iodine-Labeled Plasma Proteins, 2019
The relevant information is summarized in Table 19. The prime purpose of this table is to assist planning experiments from a protein composition point of view. Data were therefore included on a varied selection of proteins with relevance to different research areas (hem- atology, immunology, nutrition, etc.). However, no attempt was made to be comprehensive. The term, “plasma protein” , was applied in a broader sense (see Chapter 1, Section I) so as to accommodate some proteins of pathological significance. For many of the proteins listed, more than one set of analytical data is available in the literature. Reference was given in such cases to the more recent work because values from previous determinations are also frequently presented in the later publications for comparison. The molecular weights listed refer to those obtained by analytical ultracentrifugation, whenever such values are available. The state of cysteinyl residues in many proteins is not clear because the presence of free sulfhydryl groups has not been excluded by titration with Ellman’s reagent, or another criterion. The entry for these proteins under Cys-SH is a dash. In contrast, a zero entry signifies that a protein is known to contain no free SH group.
Clinical Detection of Exposure to Chemical Warfare Agents *
Published in Brian J. Lukey, James A. Romano, Salem Harry, Chemical Warfare Agents, 2019
Benedict R. Capacio, J. Richard Smith, Robert C. diTargiani, M. Ross Pennington, Richard K. Gordon, Julian R. Haigh, John R. Barr, Brian J. Lukey, Daniel Noort
Radioactive assays for AChE activity are very sensitive but require special handling and disposal. Although used in a research setting, radioactive assays have not been developed for general clinical laboratory or field use. These methods are based on the measurement of the production of [14C]carbon dioxide or choline or [3H]acetate from the hydrolysis of appropriately 14C- or 3H-labeled acetylcholine, respectively. The radioactive species are separated by Reinecke salt precipitation, differential extraction into organic solvent (Johnson and Russell, 1975), and ion-exchange chromatography or ion-exchange disks (Gordon et al., 1982). The advantage of radioactive methods is that they overcome the potentially high background in some biological samples from SH groups that may react with the Ellman reagent. On the other hand, there are notable disadvantages, including disposal of the radioactive waste, and the assays are not readily adaptable for kinetic analysis, since aliquots of the radioactive mixture must be removed for scintillation counting at specific time intervals.
Topical delivery of tetrahydrocurcumin lipid nanoparticles effectively inhibits skin inflammation: in vitro and in vivo study
Published in Drug Development and Industrial Pharmacy, 2018
Vandita Kakkar, Indu Pal Kaur, Amrit Pal Kaur, Komal Saini, Kamalinder K. Singh
THC was a generous gift sample from Sanat Pharmaceuticals Ltd. (New Delhi, India). Carbopol® 934, disodium hydrogen phosphate, sodium carbonate, sodium dihydrogen phosphate, and Tween 80 were obtained from Central Drug House (P) Ltd. (New Delhi, India). Compritol® 888ATO was obtained as gift sample from Panacea Biotec (Lalru, Punjab). Chloroform and diethyl ether were purchased from Sisco Research Laboratories (Mumbai, India). Ethylene diamine tetraacetic acid and triethanolamine were supplied by SD Fine Chemicals (Mumbai, India). Formalin and hydrochloric acid were obtained by Qualigens Fine Chemicals (Mumbai, India). Ellman reagent was purchased from Himedia Laboratories Ltd. (Mumbai, India). Methanol and polyethylene glycol were obtained from Fisher Scientific (Mumbai, India). Ethanol was obtained from Changshu Yanguan Chemicals (China). Phospholipon G was procured as a gift sample from Gattefosse SAS, France.
In vitro Anti-colorectal Cancer Potential of the Medicinal Mushroom Ganoderma neo-japonicum Imazeki in Hyperglycemic Condition: Impact on Oxidative Stress, Cell Cycle and Apoptosis
Published in Nutrition and Cancer, 2022
Meng-Fei Lau, Kek-Heng Chua, Vikineswary Sabaratnam, Umah Rani Kuppusamy
Ellman reagent was prepared freshly by dissolving 5,5′-dithiobis-(2-nitrobenzoic acid) (also known as DTNB) with reagent diluent (0.1 M Na2HPO4·7H2O and 1 mM EDTA, pH8) to make a final concentration of 4 mg/ml. The cell lysate (50 µl) was diluted in 40 µl reagent diluent, followed by the addition of 10 µl Ellman reagent. After an incubation period of 15 min, the absorbance was read at OD415 nm against the blank (PBS). A calibration curve was generated using reduced glutathione (GSH) standard (0–1.28 µM, reconstituted in reagent diluent pH6.5), and the free thiol level was expressed as µmol GSH equivalent/g protein.
Medicago sativa ameliorated cyclophosphamide-induced thrombocytopenia and oxidative stress in rats
Published in Toxin Reviews, 2023
Zahra Gholamnezhad, Vajihe Rouki, Ramin Rezaee, Mohammad Hossein Boskabady
Total thiol content was measured using Ellman’s reagent which reacts with the SH (sulfhydryl or thiol) groups to produce a yellow complex which has a peak absorbance at 412 nm. Fifty microliter of serum/homogenized tissue and 1 ml of EDTA buffer were mixed and the absorbance was read at 412 nm against tris-EDTA buffer alone (A1). Then, 20 μL of Ellman’s reagent was added and the absorbance was read again (A2). The absorbance of Ellman’s reagent alone was also read as blank (B). The following equation was used for the calculation of the total thiol concentration (Ellman 1959).