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Updated: Sep 24, 2025
Protocol for Isolation of Primary Human Hepatocytes and Corresponding Major Populations of Non-parenchymal Liver Cells
Published on: March 30, 2016
Isolation of human intrahepatic leukocytes for phenotypic and functional characterization by flow cytometry
Stephanie Kucykowicz1, Oliver E Amin1, Alice R Burton1
1Institute of Immunity and Transplantation, Division of Infection & Immunity, University College London, London, UK.
Insights
This study presents a scalable protocol for isolating human intrahepatic leukocytes (IHL), ensuring high viability and purity. The method allows for efficient immune cell analysis in liver tissue for research on homeostasis and disease.
Area of Science:
- Immunology
- Cell Biology
- Hepatology
Background:
- Tissue-resident immune cells are crucial for maintaining organ homeostasis and responding to disease.
- Understanding tissue-specific immunity requires robust methods for isolating relevant immune cell populations.
- Human intrahepatic leukocytes (IHL) play key roles in liver health and disease, but their isolation can be challenging.
Purpose of the Study:
- To develop and present a standardized, scalable protocol for the isolation of human intrahepatic leukocytes (IHL).
- To maximize the viability, purity, and yield of IHL obtained from liver tissue.
- To provide a guide for predicting IHL yield based on tissue weight for reproducible research.
Main Methods:
- A detailed protocol for the enzymatic and mechanical dissociation of human liver tissue.
- Optimization of cell washing and density gradient centrifugation steps.
- Assessment of isolated IHL viability, purity, and yield using flow cytometry.
Main Results:
- The protocol consistently yields high-purity and viable human intrahepatic leukocytes (IHL).
- The method is scalable according to liver tissue weight, ensuring reproducible results.
- A predictive guide for IHL yield per gram of tissue was established.
Conclusions:
- This protocol provides an efficient and reproducible method for isolating human intrahepatic leukocytes (IHL) for downstream applications.
- The optimized isolation technique supports detailed functional characterization and profiling of liver-resident immune cells.
- This resource is valuable for advancing research into liver immunity, homeostasis, and disease pathogenesis.
Abstract:
With the growing appreciation of tissue-resident immunity, studying tissue-specific immune cells contributing to both homeostasis and disease is imperative. Here, we provide a protocol for the isolation of human intrahepatic leukocytes (IHL) maximizing viability, purity, and yield. Our protocol is scalable by tissue weight, allowing for reproducible and efficient IHL liberation suitable for functional characterization, cell isolation, and profiling by flow (or mass) cytometry. Furthermore, we provide a "guide" to determine an expected IHL yield per gram of tissue processed. For complete details on the use and execution of this protocol, please refer to Stegmann et al. (2016), Pallett et al. (2017), Easom et al. (2018), Swadling et al. (2020), Pallett et al. (2020), and Zakeri et al. (2022).
