Presented herein are methods, devices and kits for the
mass spectrometric
immunoassay (MSIA) of proteins present in complex
biological fluids or extracts. Pipettor tips containing porous
solid supports that are covalently derivatized with affinity ligand and used to extract specific proteins and their variants from various
biological fluids. Nonspecifically bound compounds are rinsed from the extraction devices using a series of buffer and water rinses, after which the
wild type protein (and / or its variants) are eluted directly onto a target in preparation for analysis such as matrix-assisted
laser desorption /
ionization time-of-flight
mass spectrometry (MALDI-TOF MS).
Mass spectrometry of the eluted sample then follows with the retained proteins identified via accurate molecular
mass determination.
Protein and variant levels can be determined using quantitative methods in which the
protein / variant signals are normalized to signals of internal reference standard species (either doped into the samples prior to the MSIA analysis, or other endogenous
protein co-extracted with the target proteins) and the values compared to a working curves constructed from samples containing known concentrations of the protein or variants. Such MSIA devices, kits and methods have significant application in the fields of;
basic research and development,
proteomics, protein structural characterization,
drug discovery,
drug-target discovery,
therapeutic monitoring, clinical monitoring and diagnostics, as well as in the
high throughput screening of large populations to establish and recognize protein / variant patterns that are able to differentiate healthy from diseased states.