The cell line was derived from the hepatocellular carcinoma tissue of a 15-year-old Caucasian adolescent. The cells express alpha-fetoprotein, albumin, α-2-macroglobulin, α-1-antitrypsin, transferrin, α-1-antichymotrypsin, haptoglobin, ceruloplasmin, plasminogen, complement C4, C3 activator, fibrinogen, α-1-acid glycoprotein, α-2-HS-glycoprotein, β-lipoprotein, and retinol-binding protein. They also express insulin receptors and the receptor for insulin-like growth factor IGF II. The cells exhibit enzymatic activities of 3-hydroxy-3-methylglutaryl-CoA reductase and hepatic triglyceride lipase. To date, the presence of the HBV genome in these cells has not been demonstrated.

Key Points for HepG2 Cell Culture

1. HepG2 cells exhibit island-like growth patterns, with limited expansion capacity. Once they reach a certain size, they begin to grow in stacked layers.

2. To achieve uniform cell distribution, it is essential to ensure proper dissociation during cell digestion. Additionally, the initial seeding density should not be too low, as this may result in large blank areas.

3. If the experiment requires well-dispersed monolayer HepG2 cells, polylysine-coated flasks can be used for culture. Under these conditions, the cells will grow as a dispersed monolayer, but their morphology will differ significantly from uncoated conditions, appearing more elongated and spindle-shaped.

Important Notes for HepG2 Cells:

1. After thawing or passaging, some HepG2 cells will adhere while others remain suspended. Two days post-thawing, adherent cells will begin to grow outward. During growth, cells may proliferate over existing adherent layers, forming multiple cell strata.

2. Within the first week of cell recovery, viable cell clusters may remain suspended in the culture flask. During medium changes, do not discard these viable floating cells—they can be collected via centrifugation (125 × g) and reseeded into the flask. Removing or discarding these viable floating cells may reduce overall cell numbers, leading to growth arrest or cell death.

3. Minor variations in culture conditions, particularly pH and serum quality in the medium, can affect cell growth. During proliferation, intracellular vacuoles may appear, especially as cells approach confluence. Using high-quality, non-heat-inactivated fetal bovine serum (FBS) with low endotoxin levels can improve cell adhesion and monolayer formation. If cell growth is slow, increasing the serum concentration to 20% may help enhance proliferation.