Suspension trapping and data-independent acquisition enable high-flow liquid chromatography-tandem mass spectrometry-based amyloidosis typing in clinical laboratories

Mass spectrometry enables highly specific and multiplexed typing of amyloid plaques, and is widely used for this purpose in clinical reference laboratories. However, this technique relies on nanoflow liquid chromatography (LC), which reduces penetrance of this methodology due to high up-front and operational costs associated with nanoflow. This study introduces a tandem mass spectrometry (MS/MS) method with high-flow LC, which shortens the sample preparation workflow to < \(\:8\) hours. Suspension trapping was utilized to process laser capture microdissected amyloid plaques from 47 patient samples (10 from heart, 37 from kidney). The samples were then evaluated by LC-MS/MS with data-independent acquisition (DIA). The employment of suspension trapping and DIA allow for the use of high-flow LC, which is most commonly coupled with MS in clinical laboratories. A custom selection heuristic was developed to identify the most likely amyloidogenic protein from each plaque. A novel high-flow LC-DIA-MS/MS method was able to identify the amyloidogenic protein in ~ 96% of plaques in both training ( n = 25) and testing ( n = 22) sets. All inaccuracies ( n = 2) were caused by poor identification of immunoglobulin lambda proteins in post-treatment biopsy specimens and in dual amyloid-type plaques. Employing suspension trapping and DIA, a novel high-flow LC-DIA-MS/MS method was established for amyloidosis typing of LCM tissue sections, which shortens the sample preparation workflow to < 8 h, while simultaneously utilizing high-flow LC in the place of traditionally used nanoflow LC, enabling it to be used more widely in clinical laboratories.