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Hep-56.1D

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Cat# 400204

Size : 1cryovial

Brand : CLS Cell Lines Service

Hep-56.1D Cells

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Content: 1 cryovial

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Product number: 400204

Safety data sheet

Description	The Hep-56.1D hepatoma cell line is derived from a mouse liver tumor, specifically from the C3H/He mouse strain. This cell line is characterized by a notable mutation in the p53 gene, identified at different passages during in vitro propagation. Specifically, Hep-56.1D exhibits a C:G to G:C transversion at codon 132 of exon 5, resulting in an amino acid change from cysteine to tryptophan. This mutation was detected at passage number 17, suggesting a selective growth advantage conferred by the mutation, leading to its predominance in the cell population. The Hep-56.1D cell line displays a predominantly epithelial morphology, reflecting its hepatocytic origin. This is consistent with its intermediate filament protein profile, which includes the simple keratins K8 and K18, as well as vimentin and keratin K19 to varying degrees. The presence of these proteins confirms the hepatocytic nature of the cell line and its classification as a hepatoma line. Further analysis of Hep-56.1D using DNA fingerprinting did not reveal any major structural abnormalities, although some changes in the relative intensities of specific bands were observed with increasing passage numbers. This indicates genomic stability with some degree of variability over extended culture periods. The p53 mutation analysis and intermediate filament protein expression patterns together establish Hep-56.1D as a valuable model for studying hepatocellular carcinoma and the role of p53 mutations in liver tumorigenesis.
Organism	Mouse
Tissue	Liver
Disease	Hepatocellular carcinoma
Synonyms	HEP-56.1D, 56.1D, 56.1d

Age	Adult
Gender	Female
Morphology	Epithelial-like
Growth properties	Adherent

Citation	Hep-56.1D (Cytion catalog number 400204)
Biosafety level	1

Protein expression	Keratin 8, Keratin 18, Vimentin.
Tumorigenic	Yes, in C57BL/6J mice. In the third week, tumors will develop which are approx. 5-6 mm in diameter.
Ploidy status	Aneuploid
Mutational profile	p53mut, C:G -+ G:C transversion at codon 132 of mouse p53 exon 5, which corresponds to an amino acid change from cysteine to tryptophan.

Culture Medium	DMEM, w: 4.5 g/L Glucose, w: 4 mM L-Glutamine, w: 1.5 g/L NaHCO3, w: 1.0 mM Sodium pyruvate (Cytion article number 820300a)
Medium supplements	Supplement the medium with 10% FBS
Passaging solution	Accutase
Doubling time	25 to 30 hours
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.
Split ratio	A ratio of 1:4 to 1:8 is recommended
Seeding density	1 to 2 x 10^4 cells/cm^2 during routine culture
Fluid renewal	Every 3 to 4 days
Freezing recovery	>90% of the cells recovered from the freezing process within 24 to 48 h
Freeze medium	CM-1 (Cytion catalog number 800100) or CM-ACF (Cytion catalog number 806100)
Handling of cryopreserved cultures	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. Optionally, skip centrifugation but remove any remaining freezing medium after 24 hours. 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.
STR profile	M_18-3: 16 M_4-2: 20.3 M_6-7: 17 M_3-2: 14 M_19-2: 13 M_7-1: 26.2 M_1-1: 16 M_8-1: 16 M_2-1: 15 M_15-3: 22.3 M_6-4: 18 M_11-2: 16 M_1-2: 19 M_17-2: 15 M_12-1: 17 M_5-5: 17 M_X-1: 28 M_13-1: 17 Human D4/D8: -