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Proceedings Paper

Multi-target immunofluorescence using spectral FLIM-FRET for separation of undesirable antibody cross-labeling and autofluorescence (Conference Presentation)

Paper Abstract

To overcome limitations of indirect immunofluorescence, a new method is presented to employ the ostensible disadvantage of cross-labeling secondary antibodies by separation of the fluorescence signals via spectral FLIM-FRET. The undesirable cross-labeling among secondary antibodies leads to the generation of new characteristic FRET emission spectra including a change in the donor lifetime. We used a spectrally resolved FLIM detection system with pulse interleaved excitation. The combined spectral FLIM-FRET and pattern-matching analysis forms an excellent tool for use in indirect immunofluorescence that overcomes the undesirable effect of secondary antibody cross-labeling by assigning separate color channels to cross-labeled fluorescent antibodies.

Paper Details

Date Published: 9 March 2020
Proc. SPIE 11244, Multiphoton Microscopy in the Biomedical Sciences XX, 1124418 (9 March 2020); doi: 10.1117/12.2542413
Show Author Affiliations
Sumeet Rohilla, PicoQuant GmbH (Germany)
Charité Universitätsmedizin Berlin (Germany)
Benedikt Kraemer, PicoQuant GmbH (Germany)
Felix Koberling, PicoQuant GmbH (Germany)
Uwe Ortmann, PicoQuant GmbH (Germany)
Ingo Gregor, Georg-August-Univ. Göttingen (Germany)
Andreas C. Hocke, Charité Universitätsmedizin Berlin (Germany)
Rainer Erdmann, PicoQuant GmbH (Germany)

Published in SPIE Proceedings Vol. 11244:
Multiphoton Microscopy in the Biomedical Sciences XX
Ammasi Periasamy; Peter T. C. So; Karsten König, Editor(s)

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