Frontal affinity chromatography/maldi tandem mass spectrometry

G - Physics – 01 – N

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G01N 30/72 (2006.01)

Patent

CA 2566815

Sol-gel derived monolithic silica columns containing entrapped dihydrofolate reductase were used for frontal affinity chromatography of small molecule mixtures. The output from the column combined with a second stream containing the matrix molecule (HCCA) and was directly deposited onto a conventional MALDI plate that moved relative to the column via a computer controlled x-y stage, creating a semi-permanent record of the FAC run. The use of MALDI MS allowed for a decoupling of the FAC and MS methods allowing significantly higher ionic strength buffers to be used for FAC studies, which allowed for better retention of protein activity over multiple runs.

Des colonnes de silicium nonocristallin sol-gel contenant une dihydrofolate réductase piégée ont été utilisées dans la chromatographie d'affinité frontale de mélanges de petites molécules. Le résultat de la colonne a été combiné à un second flux contenant la molécule matricielle (HCCA) puis directement déposé sur une plaque MALDI classique déplacée par rapport à la colonne via un étage x-y contrôlé par ordinateur, ce qui a permis de créer un enregistrement semi-permanent de l'essai FAC. En outre, l'utilisation de MALDI MS a permis de séparer les procédés FAC et MS, et par conséquent, d'obtenir des tampons à force ionique nettement plus élevée devant servir dans des études par FAC et de parvenir à une meilleur rétention de l'activité protéique après de nombreux essais successifs.

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