Single Alexa Fluor 488 Fluorescence Measurement

To determine the fluorescence intensity of a single Alexa Fluor 488 fluorophore on actin, we monitored filaments sparsely labeled with Alexa Fluor 488. Mother filaments were polymerized using 0.8 μM labeled G-actin (containing 10% Alexa Fluor 568–actin and 0.1% Alexa Fluor 488–actin). Then, by testing different laser powers and acquisition times, we determined a set of illumination conditions (100% laser power, 1000 ms acquisition time) that ensure the clear detection of a single Alexa Fluor 488 fluorophore in our setup.

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Ghasemi F., Cao L., Mladenov M., Guichard B., Way M., Jégou A, & Romet-Lemonne G. (2024). Regeneration of actin filament branches from the same Arp2/3 complex. Science Advances, 10(4), eadj7681.

Publication 2024

Corresponding Organization : Centre National de la Recherche Scientifique

Other organizations : The Francis Crick Institute, Imperial College London

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Variable analysis

independent variables

Laser power
Acquisition time

dependent variables

Fluorescence intensity of a single Alexa Fluor 488 fluorophore on actin

control variables

Concentration of labeled G-actin (0.8 μM, containing 10% Alexa Fluor 568–actin and 0.1% Alexa Fluor 488–actin)

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