Currently, serum prostate-specific antigen (PSA) is used for the early detection of prostate cancer despite its low specificity in the range of 4-10 ng/mL. Because aberrant glycosylation is a fundamental characteristic of tumor genesis, the objective of this study was to investigate whether changes in PSA glycosylation may be used to improve the cancer specificity of PSA. We developed five lectin immunosorbant assays to analyze the glycosylation patterns of PSA in serum. Each assay sandwiches serum PSA between a PSA monoclonal antibody and a biotinylated lectin and then tags the biotin complex using a streptavidin SULFO TAG for electrochemiluminescence detection. Low limits of detection (0.04-1.35 ng/mL), good reproducibility (%CVs < 10%), and direct analysis of PSA glycosylation in sera suggest these assays may have a potential role in improving PSA's cancer specificity. Clinical performance was evaluated in 52 human subjects (26 cancer and 26 noncancer). ROC analysis showed that the total SNA assay (AUC = 0.71) appeared to perform better than percent free PSA (AUC = 0.54) in its diagnostic gray zone between 10 and 20% in a subset of 21 subjects. A separate study of 16 additional subjects showed similar findings.