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Isolation of CD133+ Liver Stem Cells for Clonal Expansion
Published on: October 10, 2011
Protocol for isolating CD163+ Kupffer cells from human liver resections
Armando Andres Roca Suarez1, Marie-Laure Plissonnier1, Maud Michelet1
1INSERM U1052, CNRS UMR-5286, Cancer Research Center of Lyon (CRCL), Lyon, France; University of Lyon, Université Claude-Bernard (UCBL), Lyon, France; The Lyon Hepatology Institute EVEREST, Lyon, France.
Abstract:
The liver microenvironment contains a wide variety of monocyte and macrophage populations. Here, we present a protocol for the specific isolation of liver-resident macrophages, known as Kupffer cells (KCs), from human liver resections. We describe steps for dissociating human liver tissues, separating non-parenchymal cells into fractions by a 2-phase iodixanol gradient, and positive selection of KCs based on the expression of CD163. We then provide instructions for validating the procedure by immunofluorescence to detect CD163. For complete details on the use and execution of this protocol, please refer to Roca Suarez, Plissonnier, et al.1.
Insights
This study details a method for isolating Kupffer cells (KCs), liver-resident macrophages, from human liver tissue. The protocol uses specific cell separation and identification techniques for accurate KC retrieval.
Area of Science:
- Immunology
- Cell Biology
- Hepatology
Background:
- The liver harbors diverse monocyte and macrophage populations.
- Kupffer cells (KCs) are the primary resident macrophages in the liver, crucial for hepatic immunity and homeostasis.
- Accurate isolation of KCs is essential for studying their functions and roles in liver diseases.
Purpose of the Study:
- To provide a detailed protocol for the specific isolation of Kupffer cells from human liver resections.
- To enable researchers to obtain pure KC populations for downstream analyses.
- To facilitate the study of KC biology in the context of human liver health and disease.
Main Methods:
- Human liver tissues are dissociated to single-cell suspensions.
- Non-parenchymal cells are separated using a two-phase iodixanol density gradient.
- Kupffer cells are isolated via positive selection based on CD163 expression.
- Immunofluorescence is employed for procedure validation, detecting CD163 expression.
Main Results:
- A reproducible protocol for isolating Kupffer cells from human liver tissue was established.
- The method allows for the enrichment of CD163-positive Kupffer cells.
- Validation confirms the specificity of the isolation procedure.
Conclusions:
- This protocol provides a reliable method for obtaining Kupffer cells from human liver resections.
- The isolated KCs can be used for various immunological and cellular studies.
- This technique will aid in advancing the understanding of liver macrophage biology and associated pathologies.
