Confirmation of peak assignments in capillary electrophoresis using immunoprecipitation. Application to D-aspartate measurements in neurons

J Chromatogr A. 2006 Feb 17;1106(1-2):56-60. doi: 10.1016/j.chroma.2005.09.037. Epub 2005 Sep 30.

Abstract

Capillary electrophoresis (CE) with laser-induced fluorescence (LIF) detection is a powerful tool for analysis of samples ranging from tissue extracts to single cells. However, accurate peak identification in electropherograms is challenging when complex biological samples are analyzed, as often matching a migration time between an analyte and corresponding standard may be insufficient to confirm the peak's identity. A method which combines single-step immunoprecipitation and CE-LIF analysis for investigation of the chiral amino acids in single cells and small tissue samples is demonstrated. D-Aspartate (D-Asp) has been reported in the central nervous system of the invertebrate neurobiological model Aplysia californica. In order to confirm the identity of D-Asp signal in the complex electropherograms of nerve tissue extracts and individual neurons, anti-D-Asp serum, preincubated with L-Asp conjugate, is added to the sample. This selectively binds the free D-Asp, creating an antibody-antigen complex with a migration time similar to that of antibody alone, but not that of D-Asp. The complete disappearance of the putative D-Asp peak confirms its identity and validates that there are no other detectable analytes co-migrating with D-Asp in the electropherogram.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Animals
  • Aplysia
  • D-Aspartic Acid / analysis*
  • Electrophoresis, Capillary / methods*
  • Immunoprecipitation / methods*
  • Neurons / chemistry*

Substances

  • D-Aspartic Acid