Single-cell suspensions were prepared from spleens 10 days post-immunization for EAE. Cells were restimulated with MOG35–55 for 72 h in supplemented DMEM medium containing inactivated 10% FCS (FBS 18, Biowest), 100 U/ml penicillin–streptomycin (BioConcept), 1 mM sodium pyruvate (Sigma), 50 M β-mercaptoethanol (Gibco), MEM non-essential amino acids (100×) (Gibco), MEM vitamins (100×) (Sigma), 200 mM l -glutamine, folic acid 14 mM (Sigma), 0.3 mM l -asparagine (Sigma), 0.7 mM l -arginine. For proliferation assays, cells were pulsed with 1 μCi of [3H]-thymidine (Hartmann Analytic) during the final 18 h and analysis of incorporated [3H]-thymidine was performed in a β-counter (Packard Top Count NXT Luminescence and Scintillation Counter). Secreted cytokines were measured after 48 h of culture with MOG35–55 by ELISA (Invitrogen).
Top count nxt luminescence and scintillation counter
Manufactured by Hewlett-Packard
Sourced in United States
The Top Count NXT Luminescence and Scintillation Counter is a laboratory instrument designed to measure luminescence and scintillation signals. It is capable of detecting and quantifying various types of luminescent and scintillation events.
Automatically generated - may contain errors
Lab products found in correlation
Penicillin streptomycin, by BioConcept (1 mentions)
Mem non essential amino acids 100, by Thermo Fisher Scientific (1 mentions)
L asparagine, by Merck Group (1 mentions)
3h thymidine, by Hartmann Analytic (1 mentions)
β mercaptoethanol, by Thermo Fisher Scientific (1 mentions)
L glutamine, by Merck Group (1 mentions)
Sodium pyruvate, by Merck Group (1 mentions)
Ultima gold, by PerkinElmer (1 mentions)
2 protocols using top count nxt luminescence and scintillation counter
1
EAE Immune Cell Profiling Protocol
2
Measurement of Liver Fatty Acid Oxidation
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Liver FAO levels were measured as previously described [17 (link)]. Liver tissue (50 mg) was placed in reaction buffer containing 0.2 mM palmitate (NEC075H250UC, 14C-palmitate at 1.25 μCi/mL, NEN Life Science, Boston, MA, USA). Homogenized liver samples were incubated on an orbital shaker-incubator (Vision, Daejeon, Korea) at 30°C. The FAO reaction produced 14CO2 was trapped with 1 N NaOH solution. After 2 hours of incubation, the reaction was stopped by the addition of 4 N sulfuric acid. The trapped 14CO2 solution was mixed with a liquid scintillation cocktail (Ultima Gold, PerkinElmer, Waltham, MA, USA) and measured using a Packard TopCount NXT Luminescence and Scintillation Counter (Packard, San Diego, CA, USA). Peroxisomal FAO was measured in the reaction buffer in the presence of 100 µM antimycin A and 12.5 µM rotenone.
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