Analytical characteristics of seminal fluid PSA differ from those of serum PSA.

To the Editor: The Technical Brief by Blase et al. [1] reported some troubling clinical conclusions while describing analytical differences among various immunoassays of prostate-specific antigen (PSA). Their study used samples consisting of free PSA and PSA complexes prepared in vitro. The free PSA molecule represents a very heterogeneous population, including pro-PSA, cleaved (“nicked”) PSA [2], PSA that can complex with a1-antichymotrypsin (ACT), and PSA that cannot complex with ACT but complexes with a2-macroglobulin [3]. In addition, heterogeneity in the carbohydrate part of the PSA molecule results in several isoforms, ranging from nonglycosylated to fully glycosylated. These variations of the free PSA molecule also affect its immunological characteristics, and for that reason, results of comparison studies done with mixtures of free PSA from seminal plasma do not represent the results that would be obtained with serum. PSA assays included in the study by Blase et al. [1] were: Hybritech Tandem-ERA, Tosoh AIA, and ACS: 180 PSA2. The designs of these assays differ, in that the Tandem-ERA and Tosoh assays use two monoclonal antibodies, whereas the ACS:180 PSA2 uses a combination of monoclonal and polyclonal antibodies. The terms “equimolar” and “skewed” are descriptive only of analytical characteristics in PSA assay design and do not take into consideration differences in assay calibration. In 1996 Stamey and colleagues [4] distributed to several manufacturers a set of nine control serum samples containing various proportions of free PSA. Six of these samples contained serum-derived free PSA; the remaining three contained free PSA purified from seminal fluid. These samples were tested with Tandem-R, Tosoh, and ACS:180 PSA2 assays. Results from the specimens with serum free PSA demonstrated that changes in the proportion of free PSA in patients’ sera do not cause a marked difference in the total PSA values measured by poly/monoclonaland dual-monoclonal-based assays (Table 1). Results from the second set of samples, which contained free PSA isolated from seminal fluid added to serum, demonstrated substantially different total PSA values obtained by the poly/monoclonaland dualmonoclonal-based assays (Table 1). These differences cannot be explained by the difference in the assay calibration or in the tracer antibodies. The results indicate notable differences in immunoreactivity between free PSA isolated from seminal fluid and free PSA in serum. Furthermore, because patients’ sera with .50% free PSA is extremely unusual [5], use of any purified sample containing .50% free PSA adds another degree of error to a hypothetical situation that has little relevance in clinical practice. Reports in the literature describing the equimolar response of various PSA assays may be used to refute claims made by Blase et al. [1]. The dual-monoclonal antibody-based assay, Tandem-R, demonstrated different affinity and reaction kinetics with free PSA isolated from serum and that from seminal fluid [3]. Blase et al. [1] did not explain why in the Tandem-ERA assay the ratio of PSA determined by Tandem-ERA and the PSA assigned value ranged from 1.13 to 1.76 for 100% free PSA. The Tandem-ERA value is 71% higher than the Tosoh value (1.76 vs 1.03). Stamey [6] used an experimental protocol similar to that of Blase et al. and compared the Tosoh and Tandem-R PSA assays. The response of the Tosoh assay was consistent with the changing proportion of free PSA, but the Tandem-R values decreased with increasing proportion of free PSA, especially when free PSA made up .50% of the total PSA [6]. These observations indicate that the molar response of the Tandem-ERA and Tandem-R assays, which use the identical antibody pair, may be affected by the assay architecture or reaction kinetics. Studies conducted with the poly/ monoclonal assay format have demonstrated excellent clinical correlation, showing linear increases in PSA concentration proportional to increases in tumor volume [7, 8]. This was especially evident for serum PSA changes from patients with organ-confined tumors grouped according to tumor volumes (as deter-