Purification of hepatocytes and sinusoidal endothelial cells from mouse liver perfusion

Fatima Cabral, Colton M. Miller, Katrina M. Kudrna, Blake E. Hass, Jocelyn G. Daubendiek, Brianna M. Kellar, Edward N. Harris

Research output: Contribution to journalArticle

15 Scopus citations

Abstract

This protocol demonstrates a method for obtaining high yield and viability for mouse hepatocytes and sinusoidal endothelial cells (SECs) suitable for culturing or for obtaining cell lysates. In this protocol, the portal vein is used as the site for catheterization, rather than the vena cava, as this limits contamination of other possible cell types in the final liver preparation. No special instrumentation is required throughout the procedure. A water bath is used as a source of heat to maintain the temperature of all the buffers and solutions. A standard peristaltic pump is used to drive the fluid, and a refrigerated table-top centrifuge is required for the centrifugation procedures. The only limitation of this technique is the placement of the catheter within the portal vein, which is challenging on some of the mice in the 18 - 25 g size range. An advantage of this technique is that only one vein is utilized for the perfusion and the access to the vein is quick, which minimizes ischemia and reperfusion of the liver that reduces hepatic cell viability. Another advantage to this protocol is that it is easy to distinguish live from dead hepatocytes by eyesight due to the difference in cellular density during the centrifugation steps. Cells from this protocol may be used in cell culture for any downstream application as well as processed for any biochemical assessment.

Original languageEnglish (US)
Article numbere56993
JournalJournal of Visualized Experiments
Volume2018
Issue number132
DOIs
StatePublished - Feb 12 2018

Keywords

  • Biochemistry
  • Cell purification
  • Collagenase
  • Hepatocyte
  • Issue 132
  • Liver perfusion
  • Mouse liver
  • Murine liver
  • Sinusoidal endothelial cells

ASJC Scopus subject areas

  • Neuroscience(all)
  • Chemical Engineering(all)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

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  • Cite this

    Cabral, F., Miller, C. M., Kudrna, K. M., Hass, B. E., Daubendiek, J. G., Kellar, B. M., & Harris, E. N. (2018). Purification of hepatocytes and sinusoidal endothelial cells from mouse liver perfusion. Journal of Visualized Experiments, 2018(132), [e56993]. https://doi.org/10.3791/56993