# A simple fixation for detection of endogenous fluorescent reporters in C. elegans that nearly eliminates intestinal autofluorescence

## TL;DR

This paper introduces a simple, chemical-free method to reduce intestinal autofluorescence in C. elegans, improving the detection of endogenous fluorescent reporters.

## Contribution

A novel fixation method that preserves GFP localization while nearly eliminating intestinal autofluorescence in C. elegans.

## Key findings

- The method preserves non-cytoplasmic GFP localization for months.
- Heat fixation nearly eliminates intestinal autofluorescence in C. elegans.
- Variability in non-membrane-anchored reporters suggests the need for empirical validation.

## Abstract

Gut granules are prominent cytoplasmic organelles that auto fluoresce when excited by much of the visible spectrum confounding fluorescence imaging of endogenous fluorescent reporters. We report a simple, chemical-free fixation method for

C. elegans

that preserves non-cytoplasmic GFP localization for months while nearly eliminating intestinal autofluorescence. To illustrate the utility and limitations of the method, we present representative images of live and heat-fixed worms expressing a variety of membrane and non-membrane localized GFP reporters expressed in diverse tissues, cells, and cellular organelles. We also describe our experience with various experimental parameters. The observed variability of non-membrane-anchored reporters suggests prudent adopters should empirically interrogate the signal fidelity of their specific reporters upon heat fixation.