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Nsc34

Manufactured by Cedarlane
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The NSC34 is a lab equipment product designed for use in research and scientific applications. It serves as a core functional component without further interpretation on its intended use.

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Lab products found in correlation

Dulbecco s modified eagle s medium dmem, by Thermo Fisher Scientific (1 mentions) Opti mem medium, by Thermo Fisher Scientific (1 mentions) Lipofectamine 3000, by Thermo Fisher Scientific (1 mentions) Fibroblast growth factor 2 (fgf2), by Miltenyi Biotec (1 mentions) Hek293t, by American Type Culture Collection (1 mentions) Sh sy5y, by American Type Culture Collection (1 mentions) Plvx acgfp1 n1 vector, by Takara Bio (1 mentions) Mda mb 231, by American Type Culture Collection (1 mentions) Neuro 2a, by American Type Culture Collection (1 mentions) C8 d1a, by American Type Culture Collection (1 mentions) Cellcrown inserts, by Merck Group (1 mentions)

5 protocols using nsc34

1

ALS Cell Model Establishment and Characterization

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NSC34, the motor neuronal cell line (Cedarlane Laboratories, Vancouver, Canada), is a hybrid of embryonic mouse spinal cord motor neuron and neuroblastoma cell line. It is characterized by extension of neuritis, generation of action potentials, expression of neurofilament proteins and choline acetyl-transferase, synthesis and storage of acetylcholine, and induction of twitching in co-cultured muscle cells [31 (link)]. The ALS cell model was established by stably transfection of mutant hSOD1G93A in NSC34, while the control was transfected with wild-type hSOD1 (WT) or empty puromycin lentivirus vector (EV).
SHSY5Y (human neuroblastoma), Hela (human cervical cancer), and HEK293T (human embryonic kidney) cells were obtained from the Cell Bank of Chinese Academy of Sciences.
The cells have been authenticated by short tandem repeat (STR) profiling and tested for mycoplasma contamination. The cells were cultivated in Dulbecco’s modified Eagle’s medium (DMEM, Gibco) supplemented by 10% fetal bovine serum (FBS, Natocor) and 100 U/mL of penicillin-streptomycin at 37 °C with 5% CO2.
Wang D., Liang W., Huo D., Wang H., Wang Y., Cong C., Zhang C., Yan S., Gao M., Su X., Tan X., Zhang W., Han L., Zhang D, & Feng H. (2022). SPY1 inhibits neuronal ferroptosis in amyotrophic lateral sclerosis by reducing lipid peroxidation through regulation of GCH1 and TFR1. Cell Death and Differentiation, 30(2), 369-382.
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2

Transfection and Infection Protocols for Evaluating sec-hGAA Expression

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Human hepatoma cells (HuH7, Thermo Fisher Scientific),47 mouse myoblast cells (C2, ATCC CRL-1772), and mouse neuronal cells (NSC34, Cedarlane)62 (link) were seeded in 6-well plates (5 × 105 cells/well) and transfected with plasmids encoding sec-hGAA (2 μg/well) using Lipofectamine 3000 in OPTIMEM medium (Thermo Fisher Scientific) according to the manufacturer’s instructions. A plasmid encoding EGFP under the control of the phosphoglycerate kinase (PGK) promoter (2 μg/well) was transfected as a control. 200 ng/well of the PGK-EGFP plasmid were co-transfected with sec-hGAA plasmids to normalize transfection efficiency. 72 hr after transfection, cells and conditioned media were harvested and analyzed for GAA enzyme activity and hGAA expression by western blot analyses. Early-passage cell cultures were used in the study.
Human skeletal muscle myoblasts (CSC-C3196, Creative Bioarray) were seeded on collagen-coated 12-well plates and infected with AAV9-hGAA or AAV9-EGFP vectors for 2 hr in OPTIMEM medium (Thermo Fisher Scientific) at an MOI of 2 × 105 vg/cell. After infection, cells were maintained using the Creative Bioarray SuperCult Skeletal Muscle Cell Growth Medium Kit supplemented with 10% fetal bovine serum and human fibroblast growth factor-2 (FGF-2, Miltenyi Biotec). Infection was repeated twice, every 48 hr; cells were harvested 48 hr following the second infection.
Colella P., Sellier P., Costa Verdera H., Puzzo F., van Wittenberghe L., Guerchet N., Daniele N., Gjata B., Marmier S., Charles S., Simon Sola M., Ragone I., Leborgne C., Collaud F, & Mingozzi F. (2018). AAV Gene Transfer with Tandem Promoter Design Prevents Anti-transgene Immunity and Provides Persistent Efficacy in Neonate Pompe Mice. Molecular Therapy. Methods & Clinical Development, 12, 85-101.
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3

Multimodal Cell Line Protocol

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NSC‐34 (Cedarlane), HEK 293T (ATCC) and SH‐SY5Y (ATCC) cells were used in this study. SH‐SY5Y cells were differentiated with retinoic acid prior to incubation with sEV. HEK 239A with GFP knocked‐into the AAVS1 locus using CRISPR was obtained from Ryan Russell (University of Ottawa). emGFP was knocked‐in with CRISPR technology. HEK 293T expressing GFP siRNA in a pre‐miR‐451 backbone was previously described (Reshke et al., 2020 (link)). Neonatal normal human dermal fibroblast (NHDF‐Neo) (Cedarlane, CC‐2509) were transduced with lentivirus (pLVX‐AcGFP1‐N1 vector, Clontech, 632154)
McCann J., Sosa‐Miranda C.D., Guo H., Reshke R., Savard A., Zardini Buzatto A., Taylor J.A., Li L, & Gibbings D.J. (2022). Contaminating transfection complexes can masquerade as small extracellular vesicles and impair their delivery of RNA. Journal of Extracellular Vesicles, 11(10), e12220.
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4

Cell Line Authentication and Mycoplasma Screening

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The following cell lines were used: NSC-34 (CLU140, Cedarlane), MDA-MB-231 (ATCC), C8S (CRL-2535, ATCC), C8D1A (CRL-2541, ATCC), MN-1 (gift of J. Cote, University of Ottawa), Neuro2a (CCL-131, ATCC), 293T (CRL-3216, ATCC), MSC-TERT (Cedarlane), and Human neonatal dermal fibroblasts (Lonza, CC-2509). BV2 cells were a gift of Dr. David Park (University of Ottawa). Bone marrow-derived macrophages were generated from femurs of wild-type C57/B6 mice selected by adherence and matured in GM-CSF for 1 week. Cell lines were not authenticated. Cell lines were verified to be free of mycoplasma contamination by assessing for cytoplasmic DAPI staining. All cell culture was performed at 37 °C in 5% CO2 in humidified incubators.
Reshke R., Taylor J.A., Savard A., Guo H., Rhym L.H., Kowalski P.S., Trung M.T., Campbell C., Little W., Anderson D.G, & Gibbings D. (2020). Reduction of the therapeutic dose of silencing RNA by packaging it in extracellular vesicles via a pre-microRNA backbone. Nature biomedical engineering, 4(1).
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5

In Vitro Cell Adhesion on SilkBridge

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In vitro cell tests were performed using RT4-D6P2T, a schwannoma cell line (ATCC catalog number CRL-2768) and Mouse Motor Neuron NSC-34 cell Line (Cedarlane catalog number CLU140).
SilkBridge  conduits were cut longitudinally and immersed in phosphate buffer saline (PBS). To allow cell adhesion, the longitudinally open conduits were pressed for a few minutes and then fixed to the culture plate-well through an insert (Sigma Aldrich, CellCrown  inserts, catalogue number Z74230). For control conditions, cell lines were plated on not coated glass coverslips.
Alessandrino A., Fregnan F., Biagiotti M., Muratori L., Bassani G.A., Ronchi G., Vincoli V., Pierimarchi P., Geuna S, & Freddi G. (2019). SilkBridge™: a novel biomimetic and biocompatible silk-based nerve conduit. Biomaterials science, 7(10).
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