Improving the Sensitivity of Fluorescence-Based Immunoassays by Photobleaching the Autofluorescence of Magnetic Beads

Shira Roth, Orr Hadass, Meir Cohen, Jasenka Verbarg, Jennifer Wilsey, Amos Danielli

Research output: Contribution to journalArticlepeer-review

27 Scopus citations


In fluorescence-based assays, usually a target molecule is captured using a probe conjugated to a capture surface, and then detected using a second fluorescently labeled probe. One of the most common capture surfaces is a magnetic bead. However, magnetic beads exhibit strong autofluorescence, which often overlaps with the emission of the reporter fluorescent dyes and limits the analytical performance of the assay. Here, several widely used magnetic beads are photobleached and their autofluorescence is reduced to 1% of the initial value. Their autofluorescence properties, including their photobleaching decay rates and autofluorescence spectra pre- and post-photobleaching, and the stability of the photobleaching over a period of two months are analyzed. The photobleached beads are stable over time and their surface functionality is retained. In a high-sensitivity LX-200 system using photobleached magnetic beads, human interleukin-8 is detected with a threefold improvement in detection limit and signal-to-noise ratio over results achievable with nonbleached beads. Since many contemporary immunoassays rely on magnetic beads as capture surfaces, prebleaching the beads may significantly improve the analytical performance of these assays. Moreover, nonmagnetic beads with low autofluorescence are also successfully photobleached, suggesting that photobleaching can be applied to various capture surfaces used in fluorescence-based assays.

Original languageEnglish
Article number1803751
Issue number3
StatePublished - 18 Jan 2019

Bibliographical note

Publisher Copyright:
© 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim


  • autofluorescence
  • fluorescence-based immunoassays
  • magnetic beads
  • magnetic modulation biosensing
  • photobleaching


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