NeutrAvidin Functionalization of CdSe/CdS Quantum Nanorods and Quantification of Biotin Binding Sites using Biotin-4-Fluorescein Fluorescence Quenching

Lisa G. Lippert, Jeffrey T. Hallock, Tali Dadosh, Benjamin T. Diroll, Christopher B. Murray, Yale E. Goldman

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

We developed methods to solubilize, coat, and functionalize with NeutrAvidin elongated semiconductor nanocrystals (quantum nanorods, QRs) for use in single molecule polarized fluorescence microscopy. Three different ligands were compared with regard to efficacy for attaching NeutrAvidin using the zero-length cross-linker 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide (EDC). Biotin-4-fluorescene (B4F), a fluorophore that is quenched when bound to avidin proteins, was used to quantify biotin binding activity of the NeutrAvidin coated QRs and biotin binding activity of commercially available streptavidin coated quantum dots (QDs). All three coating methods produced QRs with NeutrAvidin coating density comparable to the streptavidin coating density of the commercially available quantum dots (QDs) in the B4F assay. One type of QD available from the supplier (ITK QDs) exhibited ∼5-fold higher streptavidin surface density compared to our QRs, whereas the other type of QD (PEG QDs) had 5-fold lower density. The number of streptavidins per QD increased from ∼7 streptavidin tetramers for the smallest QDs emitting fluorescence at 525 nm (QD525) to ∼20 tetramers for larger, longer wavelength QDs (QD655, QD705, and QD800). QRs coated with NeutrAvidin using mercaptoundecanoicacid (MUA) and QDs coated with streptavidin bound to biotinylated cytoplasmic dynein in single molecule TIRF microscopy assays, whereas Poly(maleic anhydride-alt-1-ocatdecene) (PMAOD) or glutathione (GSH) QRs did not bind cytoplasmic dynein. The coating methods require optimization of conditions and concentrations to balance between substantial NeutrAvidin binding vs tendency of QRs to aggregate and degrade over time.

Original languageEnglish (US)
Pages (from-to)562-568
Number of pages7
JournalBioconjugate Chemistry
Volume27
Issue number3
DOIs
StatePublished - Mar 16 2016

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Nanotubes
Quantum Dots
Binding sites
Biotin
Nanorods
Semiconductor quantum dots
Quenching
Fluorescence
Binding Sites
Streptavidin
Cytoplasmic Dyneins
Coatings
Assays
biotin-4-fluorescein
neutravidin
Maleic Anhydrides
Molecules
Avidin
Fluorophores
Fluorescence microscopy

Cite this

NeutrAvidin Functionalization of CdSe/CdS Quantum Nanorods and Quantification of Biotin Binding Sites using Biotin-4-Fluorescein Fluorescence Quenching. / Lippert, Lisa G.; Hallock, Jeffrey T.; Dadosh, Tali; Diroll, Benjamin T.; Murray, Christopher B.; Goldman, Yale E.

In: Bioconjugate Chemistry, Vol. 27, No. 3, 16.03.2016, p. 562-568.

Research output: Contribution to journalArticle

Lippert, Lisa G. ; Hallock, Jeffrey T. ; Dadosh, Tali ; Diroll, Benjamin T. ; Murray, Christopher B. ; Goldman, Yale E. / NeutrAvidin Functionalization of CdSe/CdS Quantum Nanorods and Quantification of Biotin Binding Sites using Biotin-4-Fluorescein Fluorescence Quenching. In: Bioconjugate Chemistry. 2016 ; Vol. 27, No. 3. pp. 562-568.
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