A simple, sensitive and selective quantum-dot-based western blot method for the simultaneous detection of multiple targets from cell lysates

Kathryn L. Gilroy, Sarah A. Cumming, Andrew R Pitt

Research output: Contribution to journalArticle

Abstract

Quantum dots (Qdots) are fluorescent nanoparticles that have great potential as detection agents in biological applications. Their optical properties, including photostability and narrow, symmetrical emission bands with large Stokes shifts, and the potential for multiplexing of many different colours, give them significant advantages over traditionally used fluorescent dyes. Here, we report the straightforward generation of stable, covalent quantum dot-protein A/G bioconjugates that will be able to bind to almost any IgG antibody, and therefore can be used in many applications. An additional advantage is that the requirement for a secondary antibody is removed, simplifying experimental design. To demonstrate their use, we show their application in multiplexed western blotting. The sensitivity of Qdot conjugates is found to be superior to fluorescent dyes, and comparable to, or potentially better than, enhanced chemiluminescence. We show a true biological validation using a four-colour multiplexed western blot against a complex cell lysate background, and have significantly improved previously reported non-specific binding of the Qdots to cellular proteins.
LanguageEnglish
Pages547-54
Number of pages8
JournalAnalytical and Bioanalytical Chemistry
Volume398
Issue number1
Early online date27 Jun 2010
DOIs
Publication statusPublished - Sep 2010

Fingerprint

Quantum Dots
Semiconductor quantum dots
Western Blotting
Fluorescent Dyes
Color
Chemiluminescence
Antibodies
Staphylococcal Protein A
Luminescence
Multiplexing
Nanoparticles
Design of experiments
Research Design
Optical properties
Immunoglobulin G
Proteins

Keywords

  • Western blotting
  • fluorescent dyes
  • humans
  • immunoglobulin G
  • luminescent measurements
  • nerve tissue proteins
  • proteins
  • quantum dots
  • staphylococcal protein A

Cite this

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A simple, sensitive and selective quantum-dot-based western blot method for the simultaneous detection of multiple targets from cell lysates. / Gilroy, Kathryn L.; Cumming, Sarah A.; Pitt, Andrew R.

In: Analytical and Bioanalytical Chemistry, Vol. 398, No. 1, 09.2010, p. 547-54.

Research output: Contribution to journalArticle

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