Development of an ultrasensitive immunoassay for human glandular kallikrein with no cross-reactivity from prostate-specific antigen.
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 approximately 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 ( approximately 30 kDa); traces of hK2 complexed to alpha1-antichymotrypsin were present.


CONCLUSIONS
The limit of detection of the method for hK2 measurement described here ( approximately 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.
Black
M H
MH
Department of Pathology, Laboratory Medicine, Mount Sinai Hospital, Toronto, Ontario, Canada M5G 1X5.
Magklara
A
A
Obiezu
C V
CV
Melegos
D N
DN
Diamandis
E P
EP
D000818:Animals; D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D015153:Blotting, Western; D002850:Chromatography, Gel; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005453:Fluorescence; D006801:Humans; D007118:Immunoassay; D007610:Kallikreins / Q000032:analysis* / Q000276:immunology; D008297:Male; D051379:Mice; D017430:Prostate-Specific Antigen / Q000097:blood* / Q000276:immunology; D011468:Prostatectomy; D011471:Prostatic Neoplasms / Q000097:blood / Q000601:surgery; D011994:Recombinant Proteins / Q000276:immunology; D012016:Reference Values; D012661:Semen / Q000737:chemistry; D012680:Sensitivity and Specificity; D020840:Tissue Kallikreins
D000911:Antibodies, Monoclonal; D011994:Recombinant Proteins; D007610:Kallikreins; D020840:Tissue Kallikreins; D017430:Prostate-Specific Antigen