Unexpected photobleaching of Alexa 488 in a fixed bacterial sample during 2-photon excitation |
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Authors: | PC Peters C Thoni EJ Harry |
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Institution: |
a Institute for the Biotechnology of Infectious Diseases, University of Technology, Sydney, Australia
b Leica- Microsystems Pty Ltd, North Ryde, NSW, Australia |
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Abstract: | A sample of fixed bacterial cells was examined by immunofluorescence microscopy using an Alexa 488 conjugated secondary antibody for visualization. Excitation using visible light confirmed the expected photostability of this fluorophore; however, when using 2-photon excitation, Alexa 488 was rapidly and substantially photobleached. The unexpected instability of Alexa 488 under certain conditions may have deleterious consequences if not anticipated and accommodated in experimental protocols. |
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Keywords: | Alexa 488 bacteria immunofluorescence microscopy photobleaching 2-photon excitation |
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