Sensitive immunofluorescent staining of cells via generation of fluorescent nanoscale polymer films in response to biorecognition

Heather J. Avens, Brad J. Berron, Allison M. May, Katerina R. Voigt, Gregory J. Seedorf, Vivek Balasubramaniam, Christopher N. Bowman

Research output: Contribution to journalArticlepeer-review

20 Scopus citations


Immunofluorescent staining is central to nearly all cell-based research, yet only a few fluorescent signal amplification approaches for cell staining exist, each with distinct limitations. Here, the authors present a novel, fluorescent polymerization-based amplification (FPBA) method that is shown to enable similar signal intensities as the highly sensitive, enzyme-based tyramide signal amplification (TSA) approach. Being non-enzymatic, FPBA is not expected to suffer from nonspecific staining of endogenous enzymes, as occurs with enzyme-based approaches. FPBA employs probes labeled with photopolymerization initiators, which lead to the controlled formation of fluorescent polymer films only at targeted biorecognition sites. Nuclear pore complex proteins (NPCs; in membranes), vimentin (in filaments), and von Willebrand factor (in granules) were all successfully immunostained by FPBA. Also, FPBA was demonstrated to be capable of multicolor immunostaining of multiple antigens. To assess relative sensitivity, decreasing concentrations of anti-NPC antibody were used, indicating that both FPBA and TSA stained NPC down to a 1:100,000 dilution. Nonspecific, cytoplasmic signal resulting from NPC staining was found to be reduced up to 5.5-fold in FPBA as compared to TSA, demonstrating better signal localization with FPBA. FPBA's unique approach affords a combination of preferred attributes, including high sensitivity and specificity not otherwise available with current techniques.

Original languageEnglish
Pages (from-to)76-87
Number of pages12
JournalJournal of Histochemistry and Cytochemistry
Issue number1
StatePublished - Jan 2011


  • Immunocytochemistry
  • Immunofluorescent cell staining
  • Non-enzymatic
  • Polymerization
  • Signal amplification

ASJC Scopus subject areas

  • Anatomy
  • Histology


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