In vivo peroxynitrite is usually produced at lower rates, but for prolonged time periods. Thus, approaches to properly mimic the pathophysiological generation of peroxynitrite include infusion of peroxynitrite to biological systems, or the simultaneous generation of its precursors nitric oxide (NO) and superoxide anion (O2−), either using compounds such as 3-morpholinosydnonimine (SIN-1) or independent NO and O2− generation systems.25 (link) According to this, the compound SIN-1 (Enzo Life Science Inc., Farmingdale, NY, USA) was used in this study to generate nitrosative stress in human spermatozoa. This compound in solution and in the presence of molecular oxygen releases NO in a process associated with the generation of O2−, both of them react together and form peroxynitrite.29 (link) A previous report of our work group demonstrated the proper generation of peroxynitrite in suspensions of human spermatozoa using SIN-1.13 (link) SIN-1 solution was freshly prepared at 100 mmol l−1 as previously described.
Sin 1
Manufactured by Enzo Life Sciences
Sourced in United States
SIN-1 is a chemical compound that serves as a nitric oxide (NO) donor. It is commonly used in research applications to generate NO in biological systems.
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Lab products found in correlation
2 protocols using sin 1
1
In Vitro Peroxynitrite Generation in Spermatozoa
2
Peroxynitrite Decomposition by FeTPPS
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First, the ability of FeTPPS to effectively catalyze the decomposition of peroxynitrite was analyzed under our experimental conditions. For the in vitro generation of peroxynitrite the compound 3-morpholinosydnonimine (SIN-1; Enzo Life Science Inc., Farmingdale, NY, USA) was used, according to a previous report [12 (link)]. For the experiments, aliquots of sperm suspension at 2 × 106 mL−1 were exposed to 0.8 mmol/L of SIN-1 in HTF medium supplemented with different concentrations of FeTPPS (25, 50, and 100 µmol/L) for 4 h at 37 °C. A control of spermatozoa exposed to 0.8 mmol/L of SIN-1 and an untreated control were included.
In order to measure the peroxynitrite level, the fluorescent compound dihydrorhodamine 123 (DHR; Enzo Life Science Inc.) was used, and this had been previously used to detect peroxynitrite levels in human sperm suspensions [12 (link)]. The measurement of the mean fluorescence intensity (MFI) of DHR was made by flow cytometry, according to a previously described procedure [12 (link)].
In order to measure the peroxynitrite level, the fluorescent compound dihydrorhodamine 123 (DHR; Enzo Life Science Inc.) was used, and this had been previously used to detect peroxynitrite levels in human sperm suspensions [12 (link)]. The measurement of the mean fluorescence intensity (MFI) of DHR was made by flow cytometry, according to a previously described procedure [12 (link)].
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