Detection and Quantification of β-2-Microglobulin Using Mass Spectrometric Immunoassay

Abstract

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 glycosylated versions of the protein. These results demonstrate a new approach to the rapid and accurate detection/quantification of β2m present in biological fluids.