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Updated: Sep 13, 2025

A Fluorescence Fluctuation Spectroscopy Assay of Protein-Protein Interactions at Cell-Cell Contacts
Published on: December 1, 2018
Investigating the effects of temporal correlation on accurate molecular brightness determination
Ignacio Sallaberry1, Laura C Estrada1
1Universidad de Buenos Aires, Facultad de Ciencias Exactas y Naturales, Departamento de Física, Buenos Aires, Argentina; CONICET - Universidad de Buenos Aires, Instituto de Física de Buenos Aires (IFIBA), Buenos Aires, Argentina.
None:
Number and brightness, and photon counting histogram analyses, are powerful techniques for measuring stoichiometry in living cells. However, the accuracy of the results can be compromised by data temporal correlation, which increases with shorter acquisition times and slower molecular dynamics. Through simulations, we demonstrate that failing to account for data temporal correlation systematically underestimates the true molecular brightness. To overcome this limitation, we introduce the mixed assays to evaluate brightness analysis method, which decorrelates the data, allowing for accurate calculation of molecular brightness and achieving better precision with fewer data points.
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