Hep-66.3A Cells

Categories: Mouse cell lines Description: The Hep-66.4A hepatoma cell line is derived from a mouse liver tumor, specifically from the C57BL/6J mouse strain. This cell line is characterized by its hepatocytic origin, confirmed through intermediate filament protein analysis. Hep-66.4A expresses simple keratins K8 and K18, which are typical of normal liver cells, as well as vimentin and keratin K19 to varying degrees. These protein patterns confirm the hepatocytic nature of the cell line and its classification as a hepatoma line. The Hep-66.4A cell line displays a predominantly epithelial morphology, reflecting its origin from hepatocytes. This morphological phenotype is consistent with its protein expression profile. DNA fingerprint analysis of Hep-66.4A did not reveal any major structural abnormalities, indicating a degree of genomic stability. However, some changes in the relative intensities of specific bands were observed with increasing passage numbers, suggesting minor genomic variability over extended culture periods. Despite the absence of detectable p53 mutations in the primary mouse liver tumors, aberrations were found in some hepatoma lines during in vitro propagation. The Hep-66.4A cell line was analyzed for mutations in the p53 and c-Ha-ras genes. The absence of detectable mutations in the p53 gene in this line during early passages suggests a stable genetic background. This cell line serves as a valuable model for studying hepatocellular carcinoma, providing insights into the cellular and molecular mechanisms underlying liver tumorigenesis. Organism: Mouse Tissue: Liver Disease: Hepatocellular carcinoma Synonyms: HEP-66.3A, 66.3A Breed: C57BL/6J Age: Adult Gender: Female Morphology: Epithelial-like Growth Properties: Adherent Citation: Hep-66.3A (Cytion catalog number 400206) Biosafety Level: 1 Ncbi_ Taxid: 10090.0 Cellosaurus Accession: CVCL_5771 Protein Expression: Keratin 8, Keratin 18, Vimentin Tumorigenic: Yes, in B6C3F1 mice Mutational Profile: p53 wt Culture Medium: DMEM, w: 4.5 g/L Glucose, w: 4 mM L-Glutamine, w: 3.7 g/L NaHCO3, w: 1.0 mM Sodium pyruvate (Cytion article number 820300a) Supplements: Supplement the medium with 10% FBS Dissociation Reagent: Accutase Subculturing: Remove the old medium from the adherent cells and wash them with PBS that lacks calcium and magnesium. For T25 flasks, use 3-5 ml of PBS, and for T75 flasks, use 5-10 ml. Then, cover the cells completely with Accutase, using 1-2 ml for T25 flasks and 2.5 ml for T75 flasks. Let the cells incubate at room temperature for 8-10 minutes to detach them. After incubation, gently mix the cells with 10 ml of medium to resuspend them, then centrifuge at 300xg for 3 minutes. Discard the supernatant, resuspend the cells in fresh medium, and transfer them into new flasks that already contain fresh medium. Fluid Renewal: Every 3 to 5 days Freeze Medium: As a cryopreservation medium, use complete growth medium (including FBS) + 10% DMSO for adequate post-thaw viability, or CM-1 (Cytion catalog number 800100), which includes optimized osmoprotectants and metabolic stabilizers to enhance recovery and reduce cryo-induced stress. Thawing And Culturing Cells: Confirm that the vial remains deeply frozen upon delivery, as cells are shipped on dry ice to maintain optimal temperatures during transit. 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. 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. Perform all subsequent steps under sterile conditions in a flow hood, disinfecting the cryovial with 70% ethanol before opening. 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. Centrifuge the mixture at 300 x g for 3 minutes to separate the cells and carefully discard the supernatant containing residual freezing medium. 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. Adhere to established subculture protocols for continued growth and maintenance of the cell line, ensuring reliable experimental outcomes. Sterility: 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. Safety Precautions: When planning to store a cryovial in liquid nitrogen for future thawing, it is mandatory to adhere to stringent safety measures. Appropriate protective gloves and clothing are essential, and the use of a face mask or safety goggles is required during the transfer of frozen samples to or from the liquid nitrogen tank. This is to mitigate the risk of injury from potential cryovial explosions upon removal, which can result in the projection of sharp fragments. Disclaimer: Our cells are provided for in vitro laboratory research purposes exclusively and are not intended for clinical or diagnostic use, nor are they to be administered to humans or used for veterinary purposes. Users must adhere to all applicable guidelines and regulations for the handling and use of these cells in a research setting. Warranty: We stand by the promise of delivering products with high cell viability and robust culture performance. To achieve the best results, please make sure you follow the storage and culture instructions detailed in the product information sheet closely. Your adherence to these guidelines is key to success. Certificate Of Analysis: The certificate of analysis can be requested on the website or via email at info@cytion.com. Please indicate the lot number of your product in the email. Subject To Material Transfer Agreements: If you intend to use Cytion cell lines solely for internal research at a single research site, please complete and sign our Material Transfer Agreement (MTA) and submit it along with your order.For any commercial applications - including but not limited to fee-for-service work, quality control testing, product release, diagnostic use, or regulatory studies - please complete the Intended Use Form so we can prepare a suitable agreement tailored to your project.Please note: The MTA applies only to certain cell lines. If this notice and the MTA document appear on a product page, the agreement is applicable. For cell lines not covered by the MTA, no reference to the agreement will be shown. The MTA is not valid for customers in the Americas, China, or Taiwan. Please contact our U.S. entity to receive the appropriate agreement. Required Product 1: 820300a Required Product 3: 860015.0 Required Product 4: 830100.0