A Guide to Hepatocyte Markers

The liver is composed primarily of hepatocytes, with smaller populations of cholangiocytes, endothelial cells, and Kupffer cells. As the dominant cell type, hepatocytes perform essential functions including nutrient metabolism, detoxification of xenobiotics, and bile synthesis. Their central role in drug metabolism and liver disease continues to make this complex cell type a focus of intensive research. Advances in single-cell and spatial transcriptomic technologies have further revealed the complexity of hepatocyte zonation and heterogeneity across liver cell populations.

Hepatocyte markers are gene and protein signatures that serve as critical tools for characterizing liver cell populations. They are particularly valuable for distinguishing hepatocytes from other liver-resident cell types and for assessing cellular identity and function in primary cells and in vitro culture models. In this article, we highlight hepatocyte markers frequently reported in recent studies and discuss their relevance for understanding liver biology and experimental model systems. 

Liver zonation

Hepatocytes within the liver are organized into zones of specialization, known as functional zonation. Grouped as periportal (Zone 1), midzonal (Zone 2), and pericentral (Zone 3), this zonation is driven by varying gradients of oxygen, nutrients, hormones, and signaling molecules as influenced by hepatocyte proximity to the central or portal veins.

Periportal hepatocytes reside in a nutrient- and oxygen-rich environment and are responsible for many of the liver’s oxidative functions, including β-oxidation of fatty acids, gluconeogenesis, urea and protein synthesis, and cholesterol metabolism. Pericentral hepatocytes, located near the central vein in a low-oxygen environment, are specialized for glycolysis, glutamine synthesis, and xenobiotic metabolism. This zone is enriched for β-catenin signaling, which drives expression of downstream targets such as glutamine synthetase and cytochrome P450 enzymes, including CYP2E1. Midzonal hepatocytes occupy an intermediate environment and contribute to tissue maintenance. This region also contains the proliferative population under homeostatic conditions, with more limited proliferation occurring in periportal and pericentral hepatocytes.

Metabolic zonation, while traditionally defined by immunohistochemical staining of key markers, has been recommended to be analyzed using large-scale in situ gene expression profiling to better capture the full complexity and dynamic regulation of hepatocyte heterogeneity.

Cholangiocytes

An essential cellular component of the liver is the cholangiocyte, an epithelial cell type that lines the bile ducts. Although they make up only a small fraction of total liver cells, cholangiocytes are critical for bile modification and transport. Other resident liver cells include fibroblasts, hepatic stellate cells, sinusoidal endothelial cells, and immune cells such as Kupffer cells, which together support liver function through signaling and cell–cell interactions.

During liver development, bipotent hepatoblasts differentiate into either hepatocytes or cholangiocytes, the two principal epithelial lineages of the organ. Cholangiocyte-specific markers are frequently used to evaluate cellular composition, particularly in mixed-cell in vitro models. For instance, studies using hPSC-derived hepatic cells co-cultured in a 3D system with cholangiocyte and other supporting cell types have demonstrated enhanced hepatic maturation. These co-cultures show increased expression of hepatic genes, higher levels of albumin secretion, and elevated cytochrome P450 (CYP) activity, along with reduced alpha-fetoprotein (AFP) expression compared to a hepatocyte-only model. Notable cholangiocyte marker genes include CEACAM5, KRT19, KRT7, and MUC1. 

Markers in hepatocyte models

Beyond their role in characterizing tissue-derived and primary hepatocytes, hepatocyte markers are also essential for validating and studying a wide range of in vitro hepatocyte models. Primary human hepatocytes rapidly dedifferentiate in culture, limiting their use for long-term studies and driving the development of alternative systems. These include hepatocyte-like cells (HLCs) generated from pluripotent stem cells, as well as 3D culture platforms such as spheroids and organoids. Liver organoids can be produced from either primary hepatocytes or stem cell-derived progenitors, creating miniature systems that mimic aspects of liver structure and function.

In these models, markers often serve distinct purposes: in HLC differentiation protocols, they are used to monitor developmental trajectories and confirm acquisition of hepatocyte identity, while in spheroids and organoids, they are used to verify functional maturity and to distinguish hepatocytes from supporting non-parenchymal cell populations.

“While primary hepatocytes retain the full range of liver functions, alternatives like hepatic cell lines, iPSC-derived hepatocytes, or immortalized hepatocytes often lose key capabilities, such as drug metabolism or protein synthesis,” says Sujoy Lahiri, Ph.D., Lead Scientist in R&D at ATCC. “That’s why scientists must carefully characterize each cell type before use, to ensure they’re suitable for the study at hand and not missing critical functions found in primary hepatocytes.” 

Marker considerations

The appropriate choice of hepatocyte markers will depend heavily on the experimental context. Intracellular markers may be preferred when assessing hepatocyte identity or functional state at the protein level, while surface markers are often advantageous for cell isolation and live-cell analyses. Enzyme markers are often used for assessing hepatocyte function. 

“Surface markers are generally less definitive for hepatocyte identification, as many are shared with other hepatic or epithelial cell types. Therefore, intracellular markers are preferred for specificity,” says Dr. Lahiri. “For toxicity screening, CYP450 enzymes are essential, but the specific isoform must match the species being studied. For differentiation studies, albumin and HNF4α are reliable indicators of hepatocyte maturity. For long-term culture health and dedifferentiation tracking, advanced methods such as proteomic profiling or emerging epigenetic clocks may be used, though these are not yet standard in routine workflows.”

The effective use of primary hepatocytes will also require careful consideration of their unique biology and constraints. Unlike many other primary cell types, they lack proliferative capacity in vitro, which makes them a finite and non-renewable resource.

"Researchers must carefully plan ahead, often sourcing multiple donor lots to ensure enough cells for long-term studies," advises Dr. Lahiri. “Because each donor’s cells can behave differently, every lot needs to be tested and qualified before use, taking into account variations in age, metabolism, and other biological factors.”

Table of hepatocyte markers

The table below highlights protein markers of hepatocytes reported in recent literature for identifying and characterizing distinct subpopulations. Each marker entry includes links to relevant antibody and ELISA kit products, which are commonly employed in immunodetection assays. When used in combination, these markers provide a valuable reference for confirming hepatocyte identity, assessing functional states, or isolating hepatocyte populations in experimental models. 

References

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