A simple and reliable method of detecting variant transthyretins by multidimensional liquid chromatography coupled to electrospray ionization mass spectrometry

Transthyretin (TTR) variants cause amyloidosis. A method, originally report ed by us, of detecting the variants by high performance liquid chromatograp hy/electrospray ionization mass spectrometry (ESIMS) using materials precip itated with anti-TTR antiserum, has been successfully applied by several in stitutions. The method is simple and reliable, but some variants may not pr ecipitate with the antiserum or may precipitate in different yields compare d to normal TTR. Moreover, unidentified minor peaks were observed, which ma y have been derived from cross reactive materials. We have NOW devised a ne w procedure to overcome these problems. An onion exchange and reversed phas e liquid chromatography system and ESI mass spectrometer were connected in a tandem fashion using a 6 port valve and a protein trap cartridge. The pro file of ion peaks by the method was the same as that by MS with immunopreci pitates. The minor peaks were proved not to be derived from cross reactive materials, and the molecular species of these peaks were characterized. Thi s method is faster than immunoprecipitation method and using no antibody is a great benefit. The method can be applied widely to the study various pro teins, when antibodies are not available.