Mouse Neuro2a (N2a) cells (CCL-131; American Type Culture Collection, Manassas, VA) were plated at 80,000 cells/well in 24-well tissue culture dishes and, 24 h later, were cotransfected with 100 ng miR-29 target reporter plasmid plus 10 nM either control miR-Neg or miR-29a pre-miRNA synthetic oligos (Ambion, Austin, TX), along with 100 ng either TuD-Ctrl plasmid or TuD-29 expression plasmids, using Lipofectamine 2000 (Thermo Fisher Scientific, Waltham, MA), completed in triplicate for each treatment combination. At 48 h posttransfection, FF and R luciferase activities were measured using a GloMax Microplate Reader and Dual Luciferase Kit reagents (Promega). Briefly, culture media was removed and 200 μL Passive Lysis Buffer was pipetted into each well, and the plate placed on a shaker for 15 min at room temperature. Then, 10-μL lysate was transferred to duplicate wells of a 96-well white plate. Luminescence from FF and R was determined from 5-s reads after the injection of respective substrates to each well. The R/FF ratio was calculated and adjusted relative to the control (set to “1”), and results are expressed as the mean ± SEM (n = 4/group).
Neuro2a n2a cells
Manufactured by American Type Culture Collection
Sourced in United States
Neuro2a (N2a) cells are a mouse neuroblastoma cell line. They are a commonly used model system for neuroscience research.
Automatically generated - may contain errors
Lab products found in correlation
Lipofectamine 2000, by Thermo Fisher Scientific (2 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (2 mentions)
Glomax microplate reader, by Promega (1 mentions)
Dual luciferase kit reagents, by Promega (1 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions)
Opti mem reduced serum medium, by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Dulbecco s modified eagle media, by Thermo Fisher Scientific (1 mentions)
Protease inhibitor cocktail, by Merck Group (1 mentions)
4 protocols using neuro2a n2a cells
1
Regulation of miR-29 Target Reporter
2
Neuronal Differentiation of Neuro-2a Cells
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The Neuro-2a (N2a) cells, derived from mouse neuroblastoma, were purchased from the American Type Culture Collection (Manassas, VA, USA). N2a cells exhibit properties of neuronal stem cells and can differentiate into neuronal cells when treated with 20 μM retinoic acid (RA)1 (link),15 (link). The cells were incubated in culture dishes into Dulbecco’s modified Eagle’s medium (DMEM; Hyclone, Logan, UT, USA) with 10% fetal bovine serum (FBS; Serum Source International, Charlotte, NC, USA) and 1% penicillin/streptomycin (Gibco, Rockville, MD, USA) in a humidified 5% CO2 atmosphere at 37°C. When the cells reached 70–80% of confluency, the medium was replaced as a differentiation medium, which contained 2% FBS and 20 μM RA in DMEM for four days. Differentiated N2a cells were maintained in a humidified atmosphere of 5% CO2 at 37°C, and the differentiation medium was changed every two days.
3
Culturing PC12 and N2a Cell Lines
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Rat adrenal
pheochromocytoma PC12 cells and mouse neuroblastoma Neuro-2a
(N2a) cells were obtained from American Type Culture Collection (Manassas,
VA). PC12 cells were cultured in ATCC-formulated F-12K medium supplemented
with 15% HS, 2.5% FBS, and 1% antibiotics (100 units/mL PS). N2a cells
were maintained in DMEM/F12 medium containing 10% FBS and 1% PS. PC12
and N2a cells were incubated in a humidified atmosphere containing
5% CO2 at 37 °C. For all in vitro assays, topo inhibitors
CPT, DOX, and ETOP were dissolved in DMSO to prepare stock solutions
at a final DMSO concentration of less than 0.1%. The working solutions
were freshly diluted in the basal medium.
pheochromocytoma PC12 cells and mouse neuroblastoma Neuro-2a
(N2a) cells were obtained from American Type Culture Collection (Manassas,
VA). PC12 cells were cultured in ATCC-formulated F-12K medium supplemented
with 15% HS, 2.5% FBS, and 1% antibiotics (100 units/mL PS). N2a cells
were maintained in DMEM/F12 medium containing 10% FBS and 1% PS. PC12
and N2a cells were incubated in a humidified atmosphere containing
5% CO2 at 37 °C. For all in vitro assays, topo inhibitors
CPT, DOX, and ETOP were dissolved in DMSO to prepare stock solutions
at a final DMSO concentration of less than 0.1%. The working solutions
were freshly diluted in the basal medium.
4
Tau P301L Fibril Seeding in N2a Cells
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