Detection and quantification of β-2-microglobulin using mass spectrometric immunoassay

The use of mass spectrometric immunoassay (MSIA) in analyzing β-2-microglobulin (β₂m) present in human biological fluids (tears, saliva, plasma, and urine) is described. Pipettor tips containing porous affinity frits, derivatized with polyclonal anti-β₂m immunoglobulin, were manufactured and used to selectively isolate and concentrate β₂m from the biofluids, after which matrix-assisted laser desorption/ionization time-of-flight mass spectrometry was used to detect β₂m unambiguously at its characteristic molecular mass. The affinity tips were found rapid to use, requiring approximately 15 min per analysis, and exhibited low nonspecific binding properties that yielded essentially interference-free analyses. The β₂m MSIA was made quantitative by inclusion of an internal standard into the analysis for signal normalization. The resulting assay had a Linear dynamic range (R² = 0.983) covering a β²m concentration range of 0.010-1.0 mg/L with a standard error of approximately 5%. In application, urine samples from healthy individuals were screened and compared with sample from an individual suffering from renal infection. Results indicated an approximately 30-fold increase in β₂m levels in samples taken from the infected individual. During the screening, MSIA was able to distinguish between wild-type and glycosylated forms of β₂m, which made possible the accurate quantification of wild-type β₂m without interference from glycosyluted versions of the protein. These results demonstrate a new approach to the rapid and accurate detection/quantification of β₂m present in biological fluids.