K/G cells were washed once with PBS and centrifuged by using Legend Micro 17R centrifuge (Thermo Scientific) at 500 × g, 4°C for 2–3 min. The supernatant was aspirated and the pellet was retained. A total of 200 µl cytosol protein extraction kit (Beijing Solarbio Science & Technology Co., Ltd)/20 µl cell pellet (~2×106 cells) was added, placed on ice for 10 min after pipetting evenly and centrifuged at 12,000 g and 4°C for 10 min. The supernatant contained plasma protein. The remaining precipitate was added to 50–100 µl nuclear protein extraction kit (Beijing Solarbio Science & Technology Co., Ltd). the procedure used to extract plasma proteins was used to extract nucleoprotein; the obtained supernatant contained nucleoprotein, which was used for subsequent experiments.
Legend micro 17r centrifuge
Manufactured by Thermo Fisher Scientific
Sourced in Germany
The Legend Micro 17R centrifuge is a compact, refrigerated benchtop centrifuge designed for a variety of laboratory applications. It features a brushless motor and programmable controls for precise speed and temperature settings. The centrifuge can accommodate microtubes, PCR strips, and various other sample containers with a maximum rotor capacity of 24 x 1.5/2.0 mL tubes.
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Lab products found in correlation
5 protocols using legend micro 17r centrifuge
1
Cytosolic and Nuclear Protein Extraction
2
Coumarin-6 Loaded Polymeric Nanoparticles
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Briefly, Eudragit EPO (100 mg) and Kolliphor P407 (400 mg) were transferred to a 20-mL scintillation vial along with 9.95 ml of acetone. The contents in the scintillation vial were vigorously vortexed to obtain a clear solution (organic phase). Coumarin-6 (Acros, Organics) was dissolved in acetone at a concentration of 2 mg/mL and added to the organic phase containing polymers to obtain a final concentration of 0.01 mg/mL. The polymeric NPs were prepared as described earlier. The unencapsulated Coumarin-6 in the NPs was removed using Amicon Ultra-0.5 Centrifugal Filter Devices (Amicon Ultra 10K device). Briefly, NP dispersion (0.5 mL) was transferred to the Amicon-Ultra-0.5 device and centrifuged for 15 minutes at 14,000 rpm (Thermo Scientific, Legend Micro 17R centrifuge). The NPs were washed twice with ultrapure distilled water to get rid of free Coumarin-6. The filtrate was discarded and the concentrated Coumarin-6 encapsulated polymeric NPs (approximately 100 μL) were collected in a separate tube by inverting and centrifuging the Amicon Ultra filter for 2 minutes at 1,000 rpm. The concentrated NPs were then diluted to their original volume by adding approximately 400 μL of distilled water; the tube was covered with aluminum foil and stored until further use.
3
Encapsulation Efficiency of NCL-EPO-NPs
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NCL-EPO-NPs dispersion (0.5 ml) was transferred to the Amicon Ultra-0.5 device (3 kDa membrane; Fisher Scientific, Waltham, MA). The Amicon-Ultra centrifugal filter containing NCL-EPO-NPs was centrifuged for 15 minutes at 14,000 rpm (Thermo Scientific, Legend Micro 17R centrifuge). The filtrate was then diluted 10-fold with methanol, and NCL concentration in the filtrate was measured at 340 nm using a UV-Vis spectrophotometer. The % encapsulation efficiency (EE) was calculated by the following equation:
where “Dinitial” is the amount of NCL/mL of EPO-NCL-NPs dispersion and Dfree is the amount of NCL/mL of the filtrate obtained by the centrifugation of NCL-EPO-NPs.
where “Dinitial” is the amount of NCL/mL of EPO-NCL-NPs dispersion and Dfree is the amount of NCL/mL of the filtrate obtained by the centrifugation of NCL-EPO-NPs.
4
Preparation of Rhodamine-6G Encapsulated Polymeric Nanoparticles
5
Poultry Immune Response to Multivalent Vaccine
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All birds were immunized, with combined vaccine of Newcastle diseases (ND); infectious bronchitis (IB) and egg drop syndrome vaccine, inactivated (strain La Sota+Strain M41+ Strain HSH23, Beijing Ceva Huadu Biological Co, Ltd, Beijing, China) at day 140 via wing web (0.5 mL/bird). A blood sample was obtained from one bird of each replicated pen (7 birds per experimental group) via wing vein on day 147, day 154, day 161, and day 203. On the respective day of blood sample collection, serum was obtained by centrifuged at 3,000×g for 20 min at 4°C (Legend Micro 17R centrifuge, Thermo Fisher, Am Kalkberg, Germany) and was stored at −80°C until measuring antibody titers, serum immunoglobulin and serum cytokine concentration. Commercial enzyme-linked immune-sorbent assay kits (Shanghai Jianglai industrial Ltd, Shanghai, China) were used to analyze ND and IB antibody titers. The serum immunoglobulin A (sIgA), immunoglobulin G (IgG), interleukin-2 (IL-2), IL-4, IL-6, and tumor necrotic factor-α (TNF-α) were measured using chicken specific sIgA, IgG, IL-2, IL-4, IL-6, TNF-α ELISA Quantitation Kits (Shang Hai Lengton Biosicences Co. Ltd, Shang Hai, China) according to the instructions of the manufacturer and absorbance were measured at 450 nm.
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