Fluorescent Reporter-Based Paralysis Assay: A Technique to Assess Age-Associated Progressive Formation of Polyglutamine Fluorescent Reporter and Associated Paralysis in Caenorhabditis elegans
Transcription
To measure decline in proteostasis in muscle tissue, pick 20 animals, and mount them on a microscope slide with a 3% agarose pad and a 5-microliter drop of 10-millimolar sodium azide.
After all worms are immobilized, image the whole bodies of the animals using a 10x magnification lens. Use a FITC or YFP filter, and the same exposure, for every animal. When finished, discard the slides.
Count the number of foci in the body wall muscles of the whole animal. Foci are brighter, punctuated signals that can be differentiated from the dimmer soluble signal in the background.
On scoring days, look at the plates with the animals, and record the number of paralyzed animals. Then, remove paralyzed animals from the plate.
At the completion of the experiment, calculate the paralysis rate for each condition, and plot the paralysis progression.
