Genotyping of Apolipoprotein E by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Abstract

The genotyping of the various isoforms of Apolipoprotein E (apo E) has been performed using matrix-assisted laser desorption/ionization (MALDI-MS). The polymerase chain reaction was used to amplify the specific apo E gene sequence followed by digestion with Cfo I ( Clostridium formicoaceticum ), for generating restriction fragments for rapid and accurate mass analysis. An exonuclease I digestion step was introduced to remove the unused primers after PCR, which can otherwise interfere in the mass spectral analysis. By replacing the gel electrophoresis detection step with MALDI-MS, restriction isotyping of the apo E gene was achieved. Genotyping of an unknown sample obtained from an independent diagnostic laboratory demonstrated the validity of the MALDI-MS method for the routine clinical analysis of apo E. © 1998 John Wiley & Sons, Ltd.