On-Chip Mass Spectrometry-Based Immunoassay as a Tool for the Detection of Molecular Species from Prostate-Specific Antigen in Female Serum

Abstract

ABSTRACTIn females, low levels of prostate-specific antigen have been detected in hormonally dependent tissues as well as in serum and body fluids using immunometric assays. Enzymatically inactive free prostate-specific antigen was found to be the predominant form in sera of women with breast cancer, in contrast to sera from healthy women, but its molecular nature is not understood. In comparison to conventional immunoassays, mass spectrometry-based immunoassays are advantageous in terms of providing analytical data on both concentration and structural information regarding the analyte. This study was aimed at exploiting on-chip mass spectrometry-based immunoassay for profiling the molecular species of prostate-specific antigen in sera of women with breast cancer and healthy females to resolve presumed cancer-associated free prostate-specific antigen heterogeneity. Serum was analyzed using an immobilized anti-free prostate-specific antigen antibody array. Immunoreactive species were detected using surface-enhanced laser desorption–ionization time-of-flight mass spectrometry. The results obtained indicated similarities in profiles of the examined female sera regarding glycosylated full-length molecular species at 28 kDa, while noticeable differences were observed between the groups regarding immunoreactive species, reflecting modification of free prostate-specific antigen by proteolytic cleavage. The free prostate-specific antigen profile in breast cancer was specific for species at 12, 16, 21–23 kDa, and at 29 kDa, which might differ in glycosylation status. The results on low-abundance free prostate-specific antigen in female sera as yet unexploited biomarker direct further efforts to innovative use of conventional proteomic methods in diagnostic bioanalytical challenges.