Source: Hankeova, S. et al. DUCT: Double Resin Casting followed by Micro-Computed Tomography for 3D Liver Analysis. J. Vis. Exp. (2021)
This video describes the procedure for biliary system resin casting, for understanding 3D biliary architecture. This technique provides insights into the internal diameter and network connectivity of the biliary system, which could be used for liver regeneration analysis.
All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.
1. Double resin injections
2. Resin injection – Biliary system resin casting
Figure 1: Injection set for resin casting. (A) Injection set #1 comprises a 30 G needle and PE10 tubing that is ~30 cm long. Injection set #2 is composed of a 23 G needle and PE50 tubing ~30 cm long. (B) The tip of the tubing is stretched and cut at an angle to create a beveled tip. The ruler in A and B is a centimeter ruler, with major increments of 1 cm, intermediate increments of 5 mm, and minor increments of 1 mm.
Figure 2: Double resin casting flow chart. (A) Schematic showing the murine venous circulatory system and the heart with highlighted right atrium, which should be cut away before perfusion, and the left ventricle into which the needle should be inserted for perfusion to wash away the blood from the circulatory system. The inferior vena cava should be severed under the kidneys to relieve vascular pressure. (B) Bile duct resin injection flow chart. (i) Zoom image of (A) depicting where to sever the IVC. (ii) Image depicting the common bile duct (yellow dotted line) from the liver hilar region to the sphincter of Oddi (black arrowheads), with suture thread under the cleared common bile duct. (iii) Suitable position for loose overhand knot around common bile duct. (iv) The yellow dotted line and black arrowheads label the sphincter of Oddi, demonstrating the oblique angle for incision and how the opening should appear after the oblique angle incision. (v) Schematic demonstrating the orientation of the PE10 tubing bevel opening (upwards) upon insertion. (vi) Appearance of yellow resin being injected; resin should easily pass the loosely tied knot. IVC, inferior vena cava; CBD, common bile duct. (C) Portal vein resin injection flow chart. (i) The green dotted line marks the portal vein from the hilar region. The blue arrowheads label the overfilled biliary system. (ii) Suitable location for loose overhand knot around portal vein. (iii) Schematic demonstrating the bevel opening (upward) upon insertion. (iv) Appearance of liver upon injection of green resin and yellow resin; note the resin-filled blood vessel in the liver periphery. PV, portal vein. Figure 2A was created with Biorender.com.
The authors have nothing to disclose.
1.5 mL SafeSeal micro tubes | Sarstedt | 72.706 | |
23 G butterfly needle with tubing | BD bioscience | 367283 | |
23 G needle | BD bioscience | 305892 | |
30 G needle | BD bioscience | 305106 | |
Agarose | Top-Bio | P045 | |
Benzyl alcohol | Sigma Aldrich | 108006 | |
Benzyl benzoate | Sigma Aldrich | B6630 | |
Corning 50 mL tubes | Sigma Aldrich | CLS430829-500EA | polypropylene |
Cotton swabs | Medicarier | 60406 | |
Dissection Microscope | Leica Camera AG | Leica M60 | |
Dulbecco's phosphate-buffered saline | ThermoFisher Scientific | 14190144 | |
Ethanol 70% | VWR | 83801.41 | |
Falcon tube 15 mL | Verkon | 331.850.084.006 | |
Forceps curved | Fine Science Tools | 11051-10 | Fine Graefe 10 cm curved |
Forceps straight | Fine Science Tools | 11050-10 | Fine Graefe 10 cm straight |
Formaldehyde solution | Sigma Aldrich | F8775 | |
Hanks' Balanced Salt Solution | ThermoFisher Scientific | 14025092 | |
Heparin | Leo Pharma | B01AB01 | 5000 IE/mL |
Isolfurane | Baxter | FDG9623 | |
Methanol | ThermoFisher Scientific | 11413413 | |
MICROFIL | Flowtech | MV-122 | synthetic resin yellow |
MICROFIL | Flowtech | MV-120 | synthetic resin blue |
MICROFIL | Flowtech | MV-diluent | clear resin diluent |
Pasteur pipette | Verkon | 130.690.424.503 | |
Peristaltic pump | AgnThos | 010.6131.M20 | |
Rocker | VWR | 444-0142 | |
Silk suture | AgnThos | 14757 | Black silk, 4-0, sterile, 100 m |
Skin scissor | Fine Science Tools | 14058-09 | Iris straight tip 9 cm |
Spring scissor | Fine Science Tools | 15000-03 | Vannas micro, straight tip 2 mm |
Syringe 1 mL Luer | BD bioscience | 303172 | |
Tubing PE10 | BD bioscience | 427401 | |
Tubing PE50 | BD bioscience | 427411 |