After cervical dislocation, the brain and liver were removed and immediately placed on ice. After weighing, organs were homogenized in three volumes (relative to organ weight) of buffer (pH 7.6), which was prepared using 50 mM Tris-HCl, 250 mM sucrose, 60 mM KCl, 5 mM MgCl2, and 10 mM 2-mercaptoethanol. The obtained homogenate was centrifuged at 15,000× g for 15 min with a centrifuge Beckman J2-21 (Beckman Instruments, Palo Alto, CA, USA), and resulting postmitochondrial supernatant was used for the measurement of enzymatic activity in the tissue.
Beckman j2 21
Manufactured by Beckman Coulter
Sourced in United States
The Beckman J2-21 is a high-speed centrifuge designed for a wide range of laboratory applications. It features a robust and reliable design to deliver consistent performance. The J2-21 accommodates a variety of rotors and can achieve centrifugal forces up to 30,000 x g.
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Lab products found in correlation
6 protocols using beckman j2 21
1
Tissue Homogenization and Fractionation
2
Organ Isolation and Homogenization
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Following cervical dislocation of the animal, the brain and liver were removed, washed, and immediately cooled on ice. The organs were carefully weighed and homogenized in three volumes (relative to organ weight) of buffer (50 mM Tris-HCl, pH 7.6; 250 mM sucrose; 60 mM KCl; 5 mM MgCl2; 10 mM 2-mercaptoethanol). The homogenate was centrifuged at 15,000× g for 15 min centrifuge Beckman J2-21 (Beckman Instruments, Palo Alto, CA, USA)), and then postmitochondrial supernatant was used for the measurement of enzymatic activity in the organ tissues.
3
Kinnow Mandarin Peel Extraction Optimization
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Kinnow mandarin peel powders were subjected to extraction through the maceration technique as described by Elfalleh et al [31 ] with slight modifications. Preliminary studies were performed to evaluate an optimal sample/solvent ratio (1:10, 1:15, 1:20) and extraction temperature (30°C and 40°C). After the preliminary studies, extraction was carried out using different solvents—ethanol, methanol, acetone, and ethyl acetate—at three solvent concentrations (50%, 80%, 100%) with a 1:15 sample/solvent ratio and extraction temperature of 40°C. Briefly, 5-g kinnow peel powder samples were extracted by specific solvent, concentration level, extraction temperature, and sample/solvent ratio in a shaking water bath (Tecator 1024; Tecator AB, Höganäs, Sweden) for 20 hours. The extracts were filtered through Whatman filter paper 1 and centrifuged (Beckman J2-21; Beckman Coulter, Fullerton, CA, USA) at 5000 rpm for 10 minutes. The supernatant was collected, and the solvent was evaporated with a rotary evaporator (BUCHI Rotavapor, Flawil, Switzerland) under vacuum at 45°C to obtain the extract, which was further filtered through 0.45-μm membrane filter, then collected in amber glass bottles and stored at refrigeration temperature.
4
Pulp-free Orange Juice Preparation
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Orange juice was obtained from a local retailer and centrifuged at 17,696 g for 40 min (Beckman J2‐21, Beckman Coulter) to remove pulp. The pulp‐free supernatant was then filtered through a sterile filter paper of 1.2 μm pore size (Whatman, Maidstone, UK) to prevent the interference of OJ cloud particles with bacterial detection using FCM, as previously described (Anvarian, Smith, & Overton, 2018 ; Anvarian et al., 2016 ).
5
Measuring Lipid Peroxidation via MDA
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To determine the extent of lipid peroxidation in biological samples, the content of malondialdehyde (MDA) was measured. MDA forms as a result of a reaction with TBA and is expressed in nmol/g of wet tissue weight. The organs of mice were homogenized with 9 volumes w/v of cold 1.15% KCl to obtain a 10% homogenate. Then, 0.5 mL of the homogenate was mixed with 3 mL 1% H3PO4 and 1 mL 0.6% TBA aqueous solution. The reaction mixture was heated for 45 min in a boiling water bath, and after cooling, 4 mL of n-butanol was added and mixed thoroughly. The butanol phase was separated by centrifugation (Beckman J2-21, Beckman Instruments, Palo Alto, CA, USA) and used to determine light absorbance (UV/Vis spectrophotometer LAMBDA 25, (Perkin Elmer, Waltham, MA, USA) at 535 and 520 nm [125 (link)].
6
Spectrophotometric Quantification of MDA
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The lipid peroxidation marker malondialdehyde (MDA) content in biological samples were measured as described by Uchiyama et al. using the thiobarbituric acid (TBA) test [31 (link)]. The assay is based on an MDA reaction with TBA producing red MDA-TBA2 adducts. Mice organs were homogenized in cold 1.15% KCl solution to make a 10% homogenate. Samples were prepared in centrifuging tubes containing a 3 mL 1% H3PO4, 0.5 mL tissue homogenate, and 1 mL 0.6% thiobarbituric acid aqueous solution. The prepared mixture was heated in a boiling water bath for 45 min. After the samples were cooled in ice bath for 10 min, 4 mL of n-butanol was added and mixed vigorously. The butanol phase was separated by centrifugation Beckman J2-21 (Beckman Instruments, Palo Alto, CA, USA) and absorbance was measured spectrophotometrically (UV/Vis spectrophotometer LAMBDA 25, (Perkin Elmer, Waltham, MA, USA) at 535 and 520 nm. The MDA concentration was expressed as nmol/g of wet organ weight.
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