Differentiation and detection of PDGF isomers and their receptors by tunable aptamer capillary electrophoresis

Anal Chem. 2009 Sep 15;81(18):7795-800. doi: 10.1021/ac901471w.

Abstract

Tunable aptamer capillary electrophoresis (CE) techniques were developed to enable the separation and detection of platelet derived growth factor (PDGF) isomers and their receptors. Using an aptamer that formed a stable complex with the B chain but not with the A chain of PDGF, we were able to tweak the electrophoretic mobilities of the PDGF isomers for their separation. PDGF-AB bound to a single aptamer molecule was well resolved from PDGF-BB bound to two aptamer molecules. Simultaneous determination of 50 pM of two isomers was accomplished in a single analysis. Furthermore, PDGF-AB was used as a connector to bring receptor alpha and fluorescent aptamer into a single complex molecule. As a result, the formation of a (receptor alpha)-(PDGF-AB)-(fluorescent aptamer) ternary complex enabled the detection of the receptor alpha by tunable aptamer CE. A competitive assay was developed to determine receptor beta, making use of the competition between the receptor beta and fluorescent aptamer in binding to PDGF-BB. Detection limits were 0.5 nM for PDGF receptor alpha and 3 nM for receptor beta. Determination of PDGF isomers and their receptors in diluted serum samples showed no interference from the sample matrix.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aptamers, Nucleotide / chemistry
  • Aptamers, Nucleotide / metabolism*
  • Electrophoresis, Capillary / methods*
  • Fluorescent Dyes / chemistry
  • Platelet-Derived Growth Factor / analysis*
  • Platelet-Derived Growth Factor / chemistry
  • Protein Isoforms / analysis
  • Protein Isoforms / chemistry
  • Receptors, Platelet-Derived Growth Factor / analysis*
  • Receptors, Platelet-Derived Growth Factor / blood
  • Receptors, Platelet-Derived Growth Factor / chemistry
  • Recombinant Proteins / analysis
  • Recombinant Proteins / chemistry

Substances

  • Aptamers, Nucleotide
  • Fluorescent Dyes
  • Platelet-Derived Growth Factor
  • Protein Isoforms
  • Recombinant Proteins
  • Receptors, Platelet-Derived Growth Factor