TY  - JOUR
TI  - Quantitative Colocalization Analysis in Fluorescence Microscopy
AU  - Myers, Charles James
AU  - Nan, Beiyan 
VL  - 16
IS  - 20
PY  - 2026
DA  - 2026/10/20
SP  - e5832
C1  - Bio-protocol 2026;16:e5832
DO  - 10.21769/BioProtoc.5832
UR  - https://doi.org/10.21769/BioProtoc.5832
AB  - Spatial organization of macromolecules is fundamental to cellular function, with colocalization providing key insights into molecular interactions and biological processes. However, quantification remains challenging due to diverse localization patterns and irregular sample geometries. Here, we present a protocol for analyzing colocalization between two fluorescent probes using coAnalyzer, a MATLAB-based software package. coAnalyzer features a user-friendly graphical interface and supports region of interest (ROI) selection, image merging, line scanning, signal isolation, scatterplot generation, and quantitative analysis. coAnalyzer enables colocalization analysis of any two fluorophores, regardless of the proteins or dyes involved. Its broad applicability across a wide range of organisms and sample types demonstrates the robustness, flexibility, and versatility of the platform.
KW  - Colocalization
KW  - Fluorescence microscopy
KW  - Quantification
KW  - Pearson’s correlation
KW  - Manders’ correlation
JF  - Bio-protocol
SN  - 2331-8325
PB  - Bio-protocol LLC.
BIO101  - False