Immunopeptidometric assay for enzymatically active prostate-specific antigen.
BACKGROUND
Determinations of certain forms of prostate-specific antigen (PSA) have been shown to increase the specificity for prostate cancer (PCa). One such variant, proteolytically active PSA, is a potentially useful tumor marker, but it is not specifically recognized by antibodies. Using phage display libraries, we previously identified a "family" of peptides that bind specifically to active PSA. We used these to develop an immunopeptidometric assay (IPMA) that specifically detects this form of PSA.


METHODS
Microtitration plates coated with a PSA antibody were used to capture PSA, and a PSA-binding glutathione S-transferase (GST) fusion peptide was used as a tracer. Bound tracer was detected with an antibody to GST labeled with a europium chelate. PSA isoenzymes with high and low enzymatic activity were used to study binding specificity.


RESULTS
The IPMA detected enzymatically active PSA but not internally cleaved PSA and pro-PSA, which are enzymatically inactive. The assay detected 1-10% of free PSA in serum from PCa patients.


CONCLUSIONS
Peptides identified by phage display can be used to develop assays with unique specificities for enzymatically active PSA. IPMA represents a new assay principle with wide potential utility.
Wu
Ping
P
Department of Clinical Chemistry, Helsinki University Central Hospital, FIN-00029 Helsinki, Finland. ping.wu@helsinki.fi
Zhu
Lei
L
Stenman
Ulf-Hakan
UH
Leinonen
Jari
J
D005260:Female; D015200:Fluoroimmunoassay; D005982:Glutathione Transferase / Q000235:genetics; D006801:Humans; D007527:Isoenzymes / Q000097:blood; D008297:Male; D019151:Peptide Library; D010455:Peptides / Q000097:blood; D017430:Prostate-Specific Antigen / Q000097:blood* / Q000235:genetics / Q000378:metabolism*; D011498:Protein Precursors / Q000097:blood; D011993:Recombinant Fusion Proteins
D007527:Isoenzymes; D019151:Peptide Library; D010455:Peptides; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase; D017430:Prostate-Specific Antigen