Fecal Output Protocol

Abstract

This assay is used to quantify the number of fecal pellets produced over a short period of time, which serves as a measure of colonic motility. Mice are placed into individual clear plastic beakers and the number of fecal pellets in the container are counted every 5 minutes over a 30 minute period.

Steps

Prior to Assay

Prepare clean 1 liter (~12cm x 25cm) translucent beakers- sterilize if necessary, with

aluminum foil covers. These should be autoclaved before use if collection of fecal pellets for microbiome analysis is required.

Day of Set-up

Bring mice to testing room for at least 1h 0m 0s prior to assay, to acclimate (if testing mice in the same room they are housed no adjustment period is necessary).

Day of Assay

Separate 1L beakers and cut/rip foil into lids to cover each beaker individually.

Place each mouse individually into a beaker, cover with foil, and start timer. Generally, 8-10 animals can be

handled at once before start times begin to overlap.

Do not leave animals unattended while in beakers- they will attempt to jump out.

In 0h 5m 0s intervals, gently lift each cylinder, and count the number of fecal pellets present on the beaker floor and walls. Record the number of pellets for a minimum of 0h 15m 0s, up to0h 30m 0s.

Because mice are coprophagic, some pellets may disappear (be eaten) over this experiment. To be consistent, record only the cumulative number of pellets observed. Pellets produced cannot

“decrease” so the value should either stay level or increase in each 5 minute bin over the course of this assay.

Return animals to home cage, collect fecal pellets if needed.

Analysis

Comparisons should be made between genotypes and treatment groups as to the amount of pellets produced in 0h 30m 0s. There should also be comparisons of the amount of pellets produced at each 0h 5m 0s interval.