An Ex Vivo Gut Organ Culture System to Study Host-Microbiota Interactions

Abstract

Source: Azriel, S., et al. An Intestinal Gut Organ Culture System for Analyzing Host-Microbiota Interactions. J. Vis. Exp. (2021).

In this video, we describe the methodology for setting up an ex vivo gut organ culture system to study the interactions between mouse intestinal tissue fragments and specific gut bacteria. The device consists of two pairs of input-output ports, the first connecting the intestinal tissue lumen and supplying the specific bacterial suspension into it, and the second continuously supplying media to the device's wells to maintain ex vivo tissue viability.

Protocol

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

1. Experiment preparation

1. Fabrication of the gut organ culture device (3 days)
   1. Using a 3D printer, print the reusable plastic molds for the organ culture device (the device has 6 wells, with 24 small and large holes, and for the device cover lid).
      NOTE: These plastic molds may be used for the fabrication of numerous devices.
   2. Insert the blunt-end needles (22 G & 18 G) to the appropriate position within the device mold and c....

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Device
18-Gauge Blunt Needle	Mcmaster	75165a754
22-Gauge Blunt Needle	Mcmaster	75165a758
All Purpose Adhesive Sealant 100% Silicone	DAP	688
Cubic Vacuum Desiccator VDC-21+ 2 Shelves		AAAD4021
Glass Slide 1 mm Thick	Corning	2947-75X50
Mini Incubator im-10		AAH24315K
MPC 301E Vacuum PUMP		VI-412711
Plastic Quick Turn Tube Coupling Plugs	Mcmaster	51525k121
Plastic Quick Turn Tube Coupling Sockets	Mcmaster	52525k211
Sylgard 184 Silicone Elastomer	Dow		Polydimethylsiloxane, PDMS
Tubing	Mcmaster	6516t11
Zortrax M200	Zortrax		Zortrax Z-SUITE, Autodesk Fusion 360
Zortrax M200 Materials: z-ultrat	Zortrax
Medium
B27 Supplement (50x), Serum-Free T	Thermo Fisher Scientific	17504044
Iscove's Modified Dulbecco's Medium with Phenol Red (1x)	Thermo Fisher Scientific	12440061
HEPES Buffer (1M)	Thermo Fisher Scientific	15630056
Knock-Out Serum	Thermo Fisher Scientific	10828028
N2 Supplement (100x)	Thermo Fisher Scientific	A1370701
Non-Essential Amino Acid (100x)	Thermo Fisher Scientific	11140035
Surgical Tools
Large Scissors	Aseltech	11-00-10
Sharp Forceps	F.S.T	11297-10
Silk-Braided Surgical Thread	SMI	8010G
Straight Scissors	F.S.T	14091-09
Thin Forceps	F.S.T	11051-10
Organ System
0.1 µm Filter	Life Gene
0.22 µm Filter	Life Gene
5 mL Luer-Lock Syringe	B-D	309649
Greenough Stereo Microscope	ZEISS	Stemi 305
Recirculating Precision Air Heater "CUBE"		CUBE-2-LIS
Syringe Pump	New Era Pump Systems Inc	nep-ne-1600-em