Overview
This video describes the technique of isolating the intestine to evaluate the intestinal uptake of bioactive compounds. In this sample, we will isolate the intestine from an adult zebrafish to analyze the uptake of immunostimulants.
Protocol
1. Zebrafish Intestine Dissection
- After a specified period of time post intubation (e.g., 5 h and/or 24 h), place the fish using a net into 300 mg/L ethyl 3-aminobenzoate methanesulfonate (MS-222) solution for euthanasia. Make sure the operculum stops moving and there is no tail pinch reflex. Five minutes is normally sufficient.
- Pick up the euthanized animal with a net and place it on a filter paper.
NOTE: The filter paper is very useful for removing the adhesive tissue along the intestine. - Using sharp dissection scissors, make a semi-circular incision from the anus to the operculum and open incision using fine tweezers. Cut both ends of the intestine, take out all the internal organs, and place them on the filter paper.
CAUTION: Work quickly to reduce cell metabolism and death.
NOTE: Alternatively, remove the adhesive tissue in PBS and on ice. - Separate the intestine from internal organs making sure to keep its orientation (anterior to posterior intestinal segment) and stretch it out. Usually, the anterior segment of intestine is wider than posterior segment. Take care to obtain all of the intestine when dissecting.
NOTE: The posterior end is quite fine and fragile in small fish and may break off, particularly in animals <0.7 g. - Roll the intestine on the filter paper with tweezers in order to detach the adhesive tissue from the intestine.
- Proceed to prepare the intestine for various downstream analyses
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Materials
| Name | Company | Catalog Number | Comments |
| MS-222 | Sigma Aldrich | E10521 | powder |
| 10x PBS | Sigma Aldrich | P5493 | |
| Filter paper | Filter-Lab | RM14034252 |
