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Immortalized-Mouse-Microglia-(SIM-A9)

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Immortalized-Mouse-Microglia-(SIM-A9)

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	Print Version
BioSafety Level	II
Organism	C57BL/6 mouse
Source Organ	Postnatal mouse cerebral cortices
Growth Properties	Adherent
Recommended Seeding Density	Thaw entire contents into an appropriate T25 flask as specified in the Propagation instructions.
Markers	CD68, Iba1
Applications	For Research Use Only
Immortalization Method	Spontaneous immortalization
Description	Microglia cells are resident macrophages of the brain and spinal cord and they act as the first and the main form of active immune defense in the nervous system. In addition to expressing microglial specific markers CD68 and Iba1, the spontaneously Immortalized Mouse Microglia (SIM-A9) retains responsiveness to exogenous inflammatory stimulation (LPS and β-amyloid) and the ability to secret cytokines and nitric oxide as well as phagocytose biological debris. Upon LPS and IL-4 stimulation, these cells are capable of switching between the pro-inflammatory microglial phenotype, M1, which is associated with elevated iNOS and COX-2 followed by IкB and tyrosine kinase pathways and the regenerative-supportive phenotype, M2, which is identified by an increased in Arg-1 expression, respectively. SIM-A9 exhibits key attributes of primary microglia and is useful in characterization of stimulus-triggered microglial migration, proliferation and phagocytosis.
Procedure Overview
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Propagation	Use of PriCoat? T25 Flasks (G299) or Applied Cell Extracellular Matrix (G422) is required for cell adhesion to the culture vessels. Grow cells in ECM-coated culture vessels with the following conditions. The base medium for this cell line is Prigrow IV medium available from ABM (TM004). To make the complete growth medium, add the following components to the base medium: 10% heat-inactivated fetal bovine serum, 5% donor horse serum (Thermo Fisher Scientific; Cat. no. 16050130) and Penicillin/streptomycin. Stimulation of microglia can be done by adding 2.5 ng/ml LPS, or 20 μM β-amyloid fibrils (Aβ_1-42). Atmosphere: air: 95%, CO2: 5%; Temperature: 37.0°C.
Preservation	1. Freeze Medium: Complete growth medium with 20% FBS and 10% DMSO. 2. Storage Temperature: Liquid nitrogen vapour phase.
Quality Control	1) Functional assays such as the nitrate assay was performed to assess nitric oxide production; 2) ELISA was used to determine the TNFα secretion after inflammatory stimulation; 3) Western blot and immunocytochemistry were used to confirm the expression of microglial specific markers such as CD68 and Iba1 and M1/M2 phenotype-associated protein expression (COX-2/iNOS and Arg-1) after stimulation; 4) Aβ_1-42 or E. coli-derived bioparticles uptake was used to determine phagocytic activity of SIM-A9 cells.
Disclaimer	1. The CoA for this product (provided upon request) verifies the cell-type specific gene expression via RT-PCR only. All test parameters provided in the CoA are conducted using abm's standardized culture system and procedures. The stated values may vary under the end-user's culture conditions. Please verify that the product is suitable for your studies by referencing published papers or ordering RNA (0.5 μg, Cat.# C207, $450.00) or cell lysate (100 μg, Cat.# C206, $600.00) to perform preliminary experiments, or alternatively use our Gene Expression Assay Service (Cat# C138). All sales are final. 2. We strongly recommend live cell shipments for ease of cell transfer and this option can be requested at the time of ordering. Please note that the end-user will need to evaluate the feasibility of live cell shipment by taking into account the final destination's temperature variation and its geographical location. In addition, we thoroughly test our cell lines for freeze-thaw recovery. If frozen cells were received and not recovered in your lab under the exact, specified conditions (using recommended culture vessel, media, additional supplements, and atmospheric conditions), a live cell replacement is possible at a cost (plus shipping). 3. All of abm's cell biology products are for research use ONLY and NOT for therapeutic/diagnostic applications. abm is not liable for any repercussions arising from the use of its cell biology product(s) in therapeutic/diagnostic application(s). Please contact a technical service representative for more information. 4. abm makes no warranties or representations as to the accuracy of the information on this site. Citations from literature and provided for informational purposes only. abm does not warrant that such information has been shown to be accurate.