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. 2014 Dec 14;50(96):15220-3.
doi: 10.1039/c4cc07008a.

Investigating Fluorescent Dyes in Fluorescence-Assisted Screenings

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Free PMC article

Investigating Fluorescent Dyes in Fluorescence-Assisted Screenings

Joo-Eun Jee et al. Chem Commun (Camb). .
Free PMC article

Abstract

Screening of bead-based peptide libraries against fluorescent dye-labeled target proteins was found to be significantly influenced by the dye characteristics. Commercially available red fluorescent dyes with net negative charges adversely showed strong interactions with library beads. The introduction of zwitterionic dyes significantly reduced the unwanted interactions, which sheds light upon using the right fluorescent probe for acquisition of reliable results in various fluorescence-assisted applications.

Figures

Fig. 1
Fig. 1
COPAS sorting images (X-axis: time of flight (TOF); Y-axis: fluorescence intensity, a.u.) from a) a 5-mer comprehensive peptide library itself, and from its screenings against b) AL-labeled lysine, and c) AL-labeled hCAII ([dye-labeled target] = 100 nM; Detection conditions: Excitation at 640 nm solid laser, Gain = 1, PMT = 450); Font histograms from positive peptides for d) AL-labeled lysine, and e) AL-labeled hCAII.
Fig. 2
Fig. 2
COPAS sorting images from bead-based screenings of a 5-mer comprehensive peptide library against a) DY-labeled lysine, b) ZWCy5-labeled lysine, and c) ZW-labeled lysine (Detection conditions: Excitation at 640 nm solid laser, Gain = 3, PMT = 700). The concentration of each dye-labeled target for screening was adjusted to exhibit the identical emission maximum.
Fig. 3
Fig. 3
COPAS sorting images from a) a 5-mer comprehensive peptide library itself, and its screenings against b) ZW-labeled lysine, and c) ZW-labeled hCAII ([dye-labeled target] = 100 nM; Detection conditions: Excitation at 640 nm solid laser, Gain = 3, PMT = 750); Font histograms from positive peptides for d) ZW-labeled lysine, and e) ZW-labeled hCAII.
Fig. 4
Fig. 4
a) Microscopic images of TentaGel beads appending Ac-fplsk; Confocal images of TentaGel beads modified with Ac-fplsk after incubation with b) ZW-labeled lysine, c) AL-labeled lysine, d) ZW-labeled hCAII, e) AL-labeled hCAII, and f) DY-labeled hCAII. [dye-labeled target] = 100 nM.

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