Allergic reactions are caused by an immune response to a substance that is not otherwise pathogenic. The quantity of allergen required to produce anaphylaxis varies greatly between individuals. Managing, identifying, and preventing trace allergen levels are therefore primary concerns for all food manufacturers. The most typical approaches to quantifying proteins are ELISA-based methods. However, this methodology has limitations including high variability, a limited sensitivity range, and issues with antibody specificity and availability. Previously, a method for the quantitative analysis of the Kunitz Trypsin Inhibitor (KTI) was presented wherein LC-MS/MS was used to determine the quantitative levels of KTI found in soy.  Recently, a streamlined approach to the digestion, clean-up, and analysis of proteins was developed and is presented herein.  Using this single vessel sample preparation enables both the quantitative analysis of proteins, as well as high recovery for the primary sequence coverage required to confirm the sequence of known proteins.  This poster presents both the ability to quickly perform quantitative analysis of proteins and the primary sequence analysis of known proteins, in both cases using LC-MS/MS analysis.

Development and implementation of a single vessel digestion, clean-up, and sample prep protocol for quantitation and primary sequence coverage of proteins using LC-MS/MS

248th National Meeting (2014)

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