Immortalized Rat Embryonic Fibroblasts (18IM)

Immortalized Rat Embryonic Fibroblasts (18IM) are derived from rat embryonic fibroblasts isolated from 13 to 14 day old fetuses which were transfected with a plasmid containing S18-2. Transfected cells were selected for with 0.5 mg/mL G418. 18IM cells display embryonic stem cell-like properties after immortalization and subsequent loss of mesodermal markers thereafter. Loss of contact inhibition was also observed and cells were also capable of showing anchorage-independent growth. Immunostaining showed the presence of stem cell markers, such as SSEA-1, and the loss of mesodermal markers, such as vimentin and smooth muscle actin. RT-PCR also confirmed the loss of mesodermal markers, FUT4 and vimentin, and an up-regulation of stem cell markers, Sox2 and Oct3/4. This cell line may be useful as a model for the role of S18-2 in other cell types and further studies in the binding partners of S18-2 in differentiated tissues.

Immortalization Method: Transfection of plasmid encoded with GFP-fused S18–2 (GFP-S18–2) carrying a neomycin resistance gene

BioSafety Level: II

Species: Rat

Source Organ: Embryo

Growth Properties: Adherent

Morphology: Polygonal

Population Doubling: 46-56 hours

Seeding Density: 20,000 – 100,000 cells/cm²; Split Ratio: 1:2 or 1:3

Markers: SSEA-1, Sox2 and Oct3/4

Donor Age: N/A

Donor Gender: N/A

Donor Ethnicity: N/A

Propagation: The base medium for this cell line is Prigrow V medium available at abm (TM015). To make the completed growth medium, add the following components to the base medium: fetal bovine serum (TM999)* to final concentration of 5%, 0.5mg/ml Geneticin (Antibiotic G418) (G271) and Penicillin/Streptomycin Solution (G255) to a final concentration of 1%.
Carbon dioxide (CO₂): 5%, Temperature: +37.0°C.* abm does not recommend to use heat-inactivated FBS for cell culture unless specified otherwise.

Quality Control: 1) Western blot analysis for presence of S18-2 in cells; 2) Immunostaining for expression of markers ; 3) Reverse Transcription PCR for determination of presence of markers

Shipping Condition: Dry Ice

Storage Condition: liquid nitrogen or -180°C