RAG

The RAG cell line is a nonreverting 8-azaguanine-resistant mutant derived from a renal adenocarcinoma of BALB/c mice. This line was developed through alternate animal-to-tissue culture passages to enrich for the tumorigenic population while eliminating normal stromal fibroblasts. RAG cells display an ameboid to epithelioid morphology with prominent cytoplasmic processes and are resistant to hypoxanthine-guanine phosphoribosyltransferase (HGPRT)-dependent selection methods due to their enzymatic deficiency. This resistance has facilitated their use in biochemical selection systems for somatic cell hybridization experiments.

RAG cells are widely utilized as a parental line in somatic cell fusion studies due to their compatibility with fusion procedures using inactivated Sendai virus. When fused with other cell lines, such as LM(TK-) or WI-38, the hybrids retain marker chromosomes and exhibit biochemical complementation of metabolic deficiencies. These hybrids have been instrumental in mapping genetic regulatory elements and studying gene expression, particularly in kidney-associated enzymes like the ES-2 esterase. RAG hybrids provide insights into both inter- and intraspecific chromosomal segregation and functional genomics.

In addition to their role in hybridization studies, RAG cells have served as a model to study the epigenetic regulation of gene expression. Hybrid cells involving RAG often show extinction and re-expression of specific genetic traits, depending on the retention or loss of particular chromosomes. This makes the RAG cell line a valuable tool in understanding the dynamics of genetic regulation and chromosomal stability in tumorigenic cells.

37°C, 5% CO₂, humidified atmosphere.

1. Confirm that the vial remains deeply frozen upon delivery, as cells are shipped on dry ice to maintain optimal temperatures during transit.

2. Upon receipt, either store the cryovial immediately at temperatures below -150°C to ensure the preservation of cellular integrity, or proceed to step 3 if immediate culturing is required.

3. For immediate culturing, swiftly thaw the vial by immersing it in a 37°C water bath with clean water and an antimicrobial agent, agitating gently for 40-60 seconds until a small ice clump remains.

4. Perform all subsequent steps under sterile conditions in a flow hood, disinfecting the cryovial with 70% ethanol before opening.

5. Carefully open the disinfected vial and transfer the cell suspension into a 15 ml centrifuge tube containing 8 ml of room-temperature culture medium, mixing gently.

6. Centrifuge the mixture at 300 x g for 3 minutes to separate the cells and carefully discard the supernatant containing residual freezing medium.

7. Gently resuspend the cell pellet in 10 ml of fresh culture medium. For adherent cells, divide the suspension between two T25 culture flasks; for suspension cultures, transfer all the medium into one T25 flask to promote effective cell interaction and growth.

8. Adhere to established subculture protocols for continued growth and maintenance of the cell line, ensuring reliable experimental outcomes.

Mycoplasma contamination is excluded using both PCR-based assays and luminescence-based mycoplasma detection methods.

To ensure there is no bacterial, fungal, or yeast contamination, cell cultures are subjected to daily visual inspections.

For long-term preservation, place vials in vapor-phase liquid nitrogen at about −150 to −196 °C. Storage at −80 °C is acceptable only as a short interim step before transfer to liquid nitrogen.

Cryopreserved cell lines are shipped on dry ice in validated, insulated packaging with sufficient refrigerant to maintain approximately −78 °C throughout transit. On receipt, inspect the container immediately and transfer vials without delay to appropriate storage.

At Cytion, we stand behind the quality and viability of our products. We guarantee that our cells will be viable at the start of culture for 60 days after shipment from our facilities. We assure that all materials conform to the specifications described in the Certificate of Analysis and/or Product Sheet.

For full details, please refer to our Warranty Statement.