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Development and validation of a quantitative ELISA for the measurement of PSA concentration
Boris Acevedo1, Yasser Perera, Miriam Ruiz
1Recombinant Antibodies Laboratories, Pharmaceutical Division, Center for Genetic Engineering and Biotechnology, Ave 31 and 58, Cubanacán, P.O. Box 6162, 10600, Havana, Cuba. boris.acevedo@cigb.edu.cu
Background:
Prostate-specific antigen (PSA) has been used for the diagnosis and follow up of prostate cancer (PCa).
Methods:
Mouse monoclonal antibodies (MAbs) were generated against human prostate-specific antigen (PSA) for the development of a sensitive total PSA (t-PSA) assay. Two MAbs, denoted CB-PSA.4 and CB-PSA.9, with affinities of 3.7 x 10(9) and 4.7 x 10(10) l/mol, respectively, were used to develop an enzyme-linked immunosorbent assay (ELISA) for quantifying serum t-PSA concentration.
Results:
The detection limit (DL) of the assay was 0.1 microg/l (n=20, mean of "zero" standard+3S.D.), and the recovery of t-PSA was 96-103%. The within-run and between-day coefficients of variation (CV) ranged from 2.1% to 3.2%, and from 2.8% to 6.3% for PSA concentrations of 10 and 1 microg/l, respectively. The equimolar detection of t-PSA and free-PSA was demonstrated by two different methods, one consisted in the comparative evaluation of a sera panel (n=9) with our enzyme-linked immunosorbent assay (ELISA) and four commercial total PSA assays and the concordance with CIS bio total PSA assay. The assay had a linear range of 0.12 to 25 microg/l.
Conclusions:
The analytical performance characteristics of our PSA ELISA suggest that it will provide clinically useful PSA results, particularly when diagnostic algorithms are used.