Development of a genotype 325-specific proCPU/TAFI ELISA.
OBJECTIVE
A Thr/Ile polymorphism at position 325 in the coding region of proCPU has been reported. Immunological assays, fully characterized (including genotype dependency), are required for the quantitation of proCPU levels.


RESULTS
We have generated a panel of monoclonal antibodies against human, plasma-derived proCPU. Two combinations exhibiting distinct reactivities were selected for measurement of proCPU in plasma. T12D11/T28G6-HRP yielded values of 10.1+/-3.1 microg/mL (mean+/-SD, n=86; normal donors), and T32F6/T9G12-HRP yielded values of 5.4+/-3.0 microg/mL. Grouping according to the 325 genotype demonstrated that T12D11/T28G6-HRP was independent to this polymorphism whereas T32F6/T9G12-HRP revealed a complete lack of reactivity with the Ile/Ile genotype (ie, 0.0+/-0.0, 4.2+/-1.7, and 7.3+/-2.9 microg/mL for the Ile/Ile, Ile/Thr, and Thr/Thr isoforms, respectively). Commercially available antigen assays appeared to be partially dependent on the 325 genotype (eg, 44+/-8.9% and 100+/-30% for the Ile/Ile and Thr/Thr isoforms, respectively).


CONCLUSIONS
Our data demonstrate that great care should be taken when evaluating proCPU antigen values as a putative causative agent or as a diagnostic risk marker for cardiovascular events.
Laboratory for Pharmaceutical Biology and Phytopharmacology, Faculty of Pharmaceutical Sciences, Katholieke Universiteit Leuven, E. Van Evenstraat 4, B-3000 Leuven, Belgium. ann.gils@pharm.kuleuven.ac.be
Ann
A
Gils
Marie-Christine
MC
Alessi
Els
E
Brouwers
Miet
M
Peeters
Pauline
P
Marx
Judith
J
Leurs
Bonno
B
Bouma
Dirk
D
Hendriks
Irène
I
Juhan-Vague
Paul J
PJ
Declerck
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000918:Antibody Specificity; D015415:Biomarkers; D025901:Carboxypeptidase B2 / Q000032:analysis* / Q000235:genetics* / Q000276:immunology; D002318:Cardiovascular Diseases / Q000453:epidemiology; D016022:Case-Control Studies; D004789:Enzyme Activation; D004797:Enzyme-Linked Immunosorbent Assay; D005838:Genotype; D006801:Humans; D007527:Isoenzymes / Q000235:genetics* / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D010446:Peptide Fragments / Q000276:immunology; D012307:Risk Factors; D013917:Thrombin / Q000378:metabolism; D018180:Thrombomodulin / Q000378:metabolism
D000911:Antibodies, Monoclonal; D015415:Biomarkers; D007527:Isoenzymes; D010446:Peptide Fragments; D018180:Thrombomodulin; D025901:Carboxypeptidase B2; D013917:Thrombin