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Clinical Chemistry 45: 790-799, 1999;
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(Clinical Chemistry. 1999;45:790-799.)
© 1999 American Association for Clinical Chemistry, Inc.

Articles

Development of an Ultrasensitive Immunoassay for Human Glandular Kallikrein with No Cross-Reactivity from Prostate-specific Antigen

Margot H. Black1,2, Angeliki Magklara1,2, Christina V. Obiezu1,2, Dimitrios N. Melegos1 and Eleftherios P. Diamandis1,2,a

1 Department of Pathology and Laboratory Medicine, Mount Sinai Hospital, Toronto, Ontario, Canada M5G 1X5.

2 Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada M5G 1L5.
a Address correspondence to this author at: Department of Pathology and Laboratory Medicine, Mount Sinai Hospital, 600 University Ave., Toronto, Ontario, Canada M5G 1X5. Fax 416-586-8628; e-mail ediamandis{at}mtsinai.on.ca

Background: Studies demonstrating that human glandular kallikrein (hK2) is increased in prostate cancer patients have prompted speculation that this marker may of use in addition to prostate-specific antigen (PSA).

Methods: An ultrasensitive hK2 sandwich immunoassay was developed, and its detection limit, cross-reactivity, analytical recovery, precision, and linearity of dilution were evaluated. hK2 was measured in seminal plasma and sera from healthy males, females, and prostatectomized patients.

Results: Our assay has an excellent detection limit (6 ng/L) and precision (>90%). Recovery studies indicated that hK2 binds to serum protease inhibitors. All sera from healthy males had measurable hK2 concentrations (median, 402 ng/L). Almost all female sera had undetectable hK2. Serum hK2 and PSA in males correlated positively (r = 0.44), but hK2 was present at concentrations ~2.5-fold lower than PSA. The PSA/hK2 ratio in male sera was 0.1–34, with a median of 2.6. In seminal plasma, this ratio was 100–500. More than 94% of immunoreactive hK2 in serum was in the free form (~30 kDa); traces of hK2 complexed to {alpha}1-antichymotrypsin were present.

Conclusions: The limit of detection of the method for hK2 measurement described here (~20-fold lower than any other reported assay for hK2) allows the generation of new clinical information. When combined with a previously described method for PSA measurement that has no cross-reactivity from hK2, this methods allows the relative proportions of hK2 and PSA in biological fluids to be measured.© 1999 American Association for Clinical Chemistry




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