Preparation of Decellularized Rat Kidney: A Procedure to Generate Acellular Vascularized Whole-organ Scaffold from a Harvested Rat Kidney

Protokół

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.

1. Kidney Harvesting and Decellularization

1. Harvest the left kidney with the abdominal aorta and inferior vena cava.
2. Ligate the ureter, thoracic aorta, superior vena cava, and branches of the abdominal aorta.
3. Keep the organ hydrated in Dulbecco's PBS (DPBS) in a 10 cm Petri dish.
4. Cannulate the abdominal aorta and inferior vena cava with a 23G catheter. To remove residual blood, connect the cannula with a peristaltic pump and wash with DPBS (500 mL) and 16 U/mL heparin for 90 min at a rate of 5 rpm at 37 °C.
5. To decellularize the kidney, perfuse the kidney with 1% Triton X-100 (1 L) for 3 h and then with 0.75% sodium dodecyl sulfate (SDS) solution (2 L) for 6 h at a constant pressure of 40 mmHg.
   NOTE: The kidney will become transparent after 8 h.
6. To remove residual SDS, perfuse the sample with 1% penicillin in distilled water (6 L) for 18 h (overnight) and then with sterile DPBS (500 mL) and 16 U/mL heparin for 90 min.

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Materiały

Name	Company	Catalog Number	Comments
Polyethelyne 50 tubing, catheter tubing 100 ft	Braintree Scientific	.023" × .038”
3-0 silk spool vascular access/ ligation in rat	Braintree Scientific	SUT-S 110
Fine scissors to cut fascia/ connective tissue	Fine Science Tools	14058-09
25 gauge inch guide needle(for vascular catheters)	Becton Dickinson	305145
Schwartz microserrefine vascular clamps	Fine Science Tools	18052-01 (straight)
5-0 silk spool vascular access/ ligation in mouse	Braintree Scientific	SUT-S 106