Whole-liver decellularization of rat liver
Scope of the method
- Animal health
- Human health
- Basic Research
- In vitro - Ex vivo
- Animal derived cells / tissues / organs
- basic research
This method describes the steps to go from a liver to a decellularized matrix. It uses mild and strong detergents to destroy cells and keep the extracellular matrix intact. This matrix can then be used for a variety of purposes, including (but not limited to) repopulation, basis for coating and basic research.
- Laminar Air Flow (LAF) ;
- Peristaltic pump ;
- Perfusion equipment.
- Published in peer reviewed journal
Pros, cons & Future potential
Versatile tool: matirx can be used for an array of things.
- Very specific surgery and very dependent on the surgeon ;
- Small differences in cannule placement can have major implications ;
- Very hard to keep sterile ;
- Time consuming ;
- Remaining SDS is lethal for cells.
This method can be modified: cannulation of vena cava and/of arteria hepatica to create an closed environment.
This method has a lot of potential, but faces a few big obstacles. If these could be removed, this method is would be a huge step forward.
References, associated documents and other information
De Kock, Joery & Ceelen, Liesbeth & De Spiegelaere, Ward & Casteleyn, Christophe & Claes, Paul & Vanhaecke, Tamara & Rogiers, Vera. (2011). Simple and quick method for whole-liver decellularization: A novel in vitro three-dimensional bioengineering tool?. Archives of toxicology. 85. 607-12. 10.1007/s00204-011-0706-1
- - Bile cannula
- - Plastic cannula
- - Sterile surgical material (forceps, scissors, ...)
- - Sterile glass petri dish
- - Sterile injection needles (3-26G3/8)
- - Sterile drape
- - Shaving equipment (electronic and/or razor)
- - Surgical suture (mersilk, 2-0)
- - Sterile glass recipients
- - Perlonfilter
- - Sterivex® filter
- - Carbogeen (5 % CO2 and 95 % air)
- - Bidest water
- - Heparin (5000 IU/ml)
- - Sedation (e.g. 87.5 mg/kg ketamine and 12.5 mg/kg xylazine
- - Krebs-Henseleit-buffer (KHB) pH = 7.4
- - Krebs-Henseleit-buffer with calcium pH = 7.4 - 70% (v/v) ethanol solution
- - 1% triton-x solution
- - 1% SDS solution
- - Sterilize the perfusion equipment with 70% (v/v) ethanol solution.
- - Rinse with bidest water.
- - Sedate the rat (e.g. 87.5 mg/kg ketamine and 12.5 mg/kg xylazine)
- - Shave the abdomen.
- - Disinfect the abdomen with 70% alcohol solution.
- - Make a U-shape incision and put the intestines outside the abdomen.
- - Put 2 surgical sutures on the bile duct, close the lower suture.
- - Make an incision in the bile duct and cannulate.
- - Close the higher suture, fixing the cannula.
- - Put 2 surgical sutures on the vena porta without closing them.
- - Put 1 surgical suture on the vena cava inferior without closing it.
- - Inject 1 ml of diluted Heparin solution (200IU/ml) in the vena saphena medialis.
- - Close the lower suture on the vena porta.
- - Make an incision in the vena porta and cannulate with the plastic cannula.
- - Close the higher suture on the vena porta.
- - Close the suture on the vena cava.
- - Excise the liver.
- - Perfuse the liver with the perfusion equipment (15min KHB, 30 ml/min).
- - The animal dies of exsanguination.
- - Perfuse the liver 1 hour with triton-X solution
- - Perfuse the liver 1 hour with SDS solution
- - Perfuse the liver 1 hour with Bidest water
Contact personPaul Claes
OrganisationsVrije Universiteit Brussel
Pharmaceutical and Pharmacological Sciences (FARM)
In Vitro Toxicology and Dermato-cosmetology