Sep 29, 2025
Measurement of protein aggregation with PROTEOSTAT
- Maria Jose Perez J.1
- 1Institut Imagine
- Team Deleidi
Protocol Citation: Maria Jose Perez J. 2025. Measurement of protein aggregation with PROTEOSTAT. protocols.io https://dx.doi.org/10.17504/protocols.io.bp2l6zrb1gqe/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: September 25, 2025
Last Modified: September 29, 2025
Protocol Integer ID: 228172
Keywords: proteostat measurement of protein aggregation, measurement of protein aggregation, protein aggregation, proteostat measurement
Funders Acknowledgements:
ASAP
Abstract
Measurement of protein aggregation with PROTEOSTAT
Troubleshooting
Fluorescence plate-based detection
Prepare the PROTEOSTAT‱ detection dye according to the manufacturer’s instructions.
Dilute the detection dye 1:2 in 1× assay buffer (ENZ-51023-KP050).
Load 2 µL of diluted detection dye into each well of a 96-well plate with black walls and a clear bottom.
For each sample, load 30 µg of total protein and dilute in assay buffer (if needed) to a final volume of 100 µL.
Incubate the plate for 15 minutes at room temperature in the dark.
Measure fluorescence at 550 nm excitation and 600 nm emission using a fluorescence plate reader (PerkinElmer Envision 2105).
Imaging-based detection
Use the PROTEOSTAT‱ Aggresome Detection Kit (Cat. No. ENZ-51035-KIT) for imaging-based detection.
Fix cells or tissue sections and permeabilize with 0.1% Triton X-100 in PBS.
Incubate samples with detection reagent for 30 minutes at room temperature in the dark, according to the manufacturer’s instructions.
Wash samples with PBS.
Counterstain samples with DAPI.
Image using a Leica TCS SP8 confocal microscope with a 60×/1.4 NA oil immersion lens.