The temporoparietal cortex (n = 5 rats in each group) was mechanically homogenized in phosphate-buffered saline solution (PBS), pH 7.4, and centrifuged at 12,500 rpm for 30 min at 4°C by using a Z 216 MK microcentrifuge (HERMLE Labortechnik; Wehingen, Germany). The production of NO was assessed through the accumulation of nitrites (NO2−) in the supernatants as described elsewhere [21 (link)]. Briefly, the nitrite concentration in 100 μL of the supernatant was measured by using a colorimetric reaction generated by the addition of 100 μL of Griess reagent, composed of equal volumes of 0.1% N-(1-naphthyl)ethylenediamine dihydrochloride and 1.32% sulfanilamide in 60% acetic acid. The absorbance of the samples was determined at 540 nm with a SmartSpec 3000 spectrophotometer (Bio-Rad; Hercules, CA, USA) and interpolated by using a standard curve of NaNO2 (1 to 10 μM) to calculate the nitrite concentration.
Z 216 mk microcentrifuge
Manufactured by Hermle
Sourced in Germany
The Z 216 MK is a microcentrifuge designed for general laboratory applications. It features a maximum speed of 15,000 rpm and a maximum relative centrifugal force (RCF) of 21,382 x g. The unit accommodates up to 18 microtubes of 1.5 or 2.0 mL capacity. The Z 216 MK includes a brushless DC motor and a maintenance-free drive system.
Automatically generated - may contain errors
Lab products found in correlation
Smartspec 3000 spectrophotometer, by Bio-Rad (1 mentions)
Uv 1800 uv vis spectrophotometer, by Shimadzu (1 mentions)
Phosphate buffered saline (pbs), by PanEco (1 mentions)
Versene, by PanEco (1 mentions)
Trypsin, by MP Biomedicals (1 mentions)
Nanodrop 1000 spectrophotometer, by Thermo Fisher Scientific (1 mentions)
Bio rs 24 mini rotator, by Biosan (1 mentions)
7 protocols using z 216 mk microcentrifuge
1
Nitrite Quantification in Rat Cortex
2
Valsartan Nanosuspension Preparation
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The valsartan-loaded nanosuspension samples were microcentrifuged at 15,000 rpm for 30–60 min with a Hermle Z216 MK Microcentrifuge (Hermle AG, Gosheim, Germany). After discarding the supernatant, the pellets were dried at 40 °C for 48 h until a constant weight was reached. The yield was calculated based on the particle mass in the entire sample, and the theoretically weighed combined mass of the polymer and drug.
3
Quantifying Hippocampal Nitric Oxide
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To investigate the role of nitric oxide, the levels of nitrites were evaluated in the hippocampus and temporoparietal cerebral cortex at the end of the study. After 34 days of reperfusion, the hippocampus and temporoparietal cortex of five rats from each group were homogenized in a phosphate-buffered saline solution with a pH of 7.4. The homogenates were centrifuged at 12,500 rpm for 30 min at 4 °C using a Z 216 MK microcentrifuge (HERMLE Labortechnik, Wehingen, Germany). The absorbance of the samples was measured at a wavelength of 540 nm with a NanoDrop (Thermo Scientific Technologies; Wilmington, DE, USA), and the production of NO was calculated by interpolating values in a standard curve of NaNO2 (1–10 μM) in the supernatants, as follows: 10 μL of the supernatant plus 10 μL of Griess reagent composed of 0.1 % N-(1-naphthyl) ethylenediamine dihydrochloride and 1.32 % of sulfanilamide in 60 % acetic acid (1:1) to produce a colorimetric reaction. Complete details of this procedure are described in Ref. [61 (link)].
4
Spectrophotometric Analysis of Valsartan Nanoparticles
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The valsartan entrapped in the nanoparticles was determined by a spectrophotometric method using a UV-1800 UV–vis spectrophotometer (Shimadzu Corporation, Kyoto, Japan). The non-encapsulated drug was measured in the supernatant after microcentrifugation with a Hermle Z216 MK Microcentrifuge (Hermle AG, Gosheim, Germany) of the nanosuspensions. Calibration was prepared at the applied concentrations of the emulsifier with a detectable linear calibration range from 2.5–40 µg/mL. In the case of samples that were formulated without PVA emulsifier, a phosphate-buffered solution at pH 6.8 was used for calibration in the same concentration range. The absorbance of the diluted sample solutions was measured at 250 nm.
5
Culturing and Preparing MCF-7 Cells for Imaging
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MCF-7 cells were cultured in a CO2 incubator for 48 h; then, the incubated cells were removed from the flask surface using a solution of 0.25% Trypsin (MP Biomedicals, Santa Ana, CA, USA)/Versene (0.2% EDTA in phosphate buffer, PanEco, Russia) (1:4). Next, the cells were pelleted and washed in 0.01M phosphate-buffered saline (PBS; PanEco, Russia) by centrifugation (at 150 G, 5 min, +37.0 ± 1.0 °C) on a Z 216 MK microcentrifuge (Hermle Labortechnik GmbH, Wehingen, Germany). After removal of the supernatant, the cell pellet was resuspended in 300 μL of PBS. A suspension of viable cells at a concentration 5 × 104 cells/mL was therewith obtained.
The samples intended for imaging were prepared by translocating the suspension of viable cells into a zone restricted by a thin layer of silicone grease on the reflecting surface of a mirror slide; then, the zone containing the cells was covered with a coverslip, whereupon the samples were immediately examined by LIM.
The samples intended for imaging were prepared by translocating the suspension of viable cells into a zone restricted by a thin layer of silicone grease on the reflecting surface of a mirror slide; then, the zone containing the cells was covered with a coverslip, whereupon the samples were immediately examined by LIM.
6
Nitric Oxide Measurement in Rat Brain
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The NO production was assessed by the accumulation of nitrites (NO2−) by the modified technique of Griess as described previously [69 (link)]. The cerebral cortex and hippocampus were obtained 1 h after the memory test (n = 5 rats in each group), mechanically homogenized in phosphate-buffered saline solution pH 7.4 (PBS), and centrifuged at 12,500 rpm for 30 min at 4°C by using a Z216MK microcentrifuge (HERMLE Labortechnik; Wehingen, Germany). Briefly, the NO concentration was measured in 10 μL of supernatant after the addition of 10 μL of Griess reagent (composed of equal volumes of 0.1% N-(1-naphthyl) ethylenediamine dihydrochloride and 1.32% sulfanilamide in 60% acetic acid). In addition, the optical density in a 2 μL sample was measured with a NanoDrop 1000 Spectrophotometer (Thermo Fisher Scientific, Bancroft Building, Wilmington, USA) at 540 nm, and the optical density values were interpolated from the standard curve of NaNO2 (1 to 10 μM) to obtain NO concentration.
7
In Vitro Drug Release Characterization of Nanocomposites
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During the pre-programmed drug release experiments fluorescein as a model compound was embedded into polymers (PVA, PVA-4, PVA-5 and PVA-8) synthesized with different polarities, from which fluorescein release was monitored by Metertech SP-8001 UV/Visible Spectrophotometer at 490 nm. (Doughty, 2010) (link) The physiological condition was ensured by phosphate buffered saline aqueous solution (50 ml) and incubation at 37 °C, continuous shaking at 200 rpm. Samples were taken at the 0 In each case, the experiments were repeated at least three times, the standard deviation of the results was well within 10%.
The sorafenib-loaded nanocomposite was subjected to biorelevant in vitro drug release experiments, that is, after their washing, 10 mg nanocomposites were re-suspended in 15 ml human blood plasma containing 0.03% sodium azide bactericide. The NPs in release medium were incubated at 37°C in G24 Environmental Incubator Shaker (New Brunswick Scientific Co. Inc., USA) and shaken by BIO RS-24 Mini-rotator (Biosan, Latvia) for 2 days at 700 rpm. At predetermined intervals, 0.5 ml of each sample was centrifuged (Hermle Z216 MK microcentrifuge, Germany) for 20 min at 15,000 rpm, washed three times and the gained nanoparticles were dissolved in 0.5 ml ethanol. The sorafenib concentration was analyzed after HPLC separation as described above.
The sorafenib-loaded nanocomposite was subjected to biorelevant in vitro drug release experiments, that is, after their washing, 10 mg nanocomposites were re-suspended in 15 ml human blood plasma containing 0.03% sodium azide bactericide. The NPs in release medium were incubated at 37°C in G24 Environmental Incubator Shaker (New Brunswick Scientific Co. Inc., USA) and shaken by BIO RS-24 Mini-rotator (Biosan, Latvia) for 2 days at 700 rpm. At predetermined intervals, 0.5 ml of each sample was centrifuged (Hermle Z216 MK microcentrifuge, Germany) for 20 min at 15,000 rpm, washed three times and the gained nanoparticles were dissolved in 0.5 ml ethanol. The sorafenib concentration was analyzed after HPLC separation as described above.
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