Bead-based ELISA for validation of ovarian cancer early detection markers.
OBJECTIVE
Efforts to validate ovarian cancer early detection biomarkers with immunoassays are challenged by the limited specimen volumes available. We sought to develop a specimen-efficient assay to measure CA125 in serum, assess its reproducibility, validity, and performance, and test its potential for multiplexing and combining with human epididymis protein 4 (HE4), a promising novel ovarian cancer marker.


METHODS
Four pairs of commercially available anti-CA125 antibodies and one pair of anti-HE4 antibodies were evaluated for accuracy in measuring known concentrations of antigen on a bead-based platform. The two best pairs were further assessed for reproducibility, validity, and the ability to discriminate between blinded serum samples obtained from ovarian cancer cases (n = 66) and women without ovarian cancer (n = 125).


RESULTS
Suitability for use in a bead-based assay varied across CA125 antibody pairs. Two CA125 bead-based assays were highly reproducible (overall correlations between replicates >/= 0.95; coefficients of variation < 0.2) and strongly correlated with the research standard CA125II RIA (correlations >/= 0.9). Their ability to distinguish ovarian cancer cases from non-cases based on receiver operating characteristic analyses (area under the curve, AUC, of 0.85 and 0.84) was close to that of the CA125II RIA (AUC, 0.87). The HE4 bead-based assay showed lower reproducibility but yielded an AUC of 0.89 in receiver operating characteristics analysis. Multiplexing was not possible but a composite marker including CA125 and HE4 achieved an AUC of 0.91.


CONCLUSIONS
Optimization procedures yielded two bead-based assays for CA125 that perform comparably to the standard CA125II RIA, which could be combined with an HE4 bead-based assay to improve diagnostic performance, and requires only 15 muL of sample each.
Scholler
Nathalie
N
Translational Outcomes Research Laboratory, Fred Hutchinson Cancer Research Center, Public Health Sciences, and Harborview Medical Center, University of Washington, Seattle, Washington, USA.
Crawford
Meghan
M
Sato
Alicia
A
Drescher
Charles W
CW
O'Briant
Kathy C
KC
Kiviat
Nancy
N
Anderson
Garnet L
GL
Urban
Nicole
N
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D014408:Biomarkers, Tumor / Q000097:blood*; D018394:CA-125 Antigen / Q000097:blood*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D030501:Epididymal Secretory Proteins / Q000032:analysis*; D005260:Female; D006801:Humans; D008875:Middle Aged; D010051:Ovarian Neoplasms / Q000097:blood / Q000175:diagnosis*; D011237:Predictive Value of Tests; D012372:ROC Curve; D012680:Sensitivity and Specificity; D023083:beta-Defensins
D014408:Biomarkers, Tumor; D018394:CA-125 Antigen; C426184:DEFB126 protein, human; D030501:Epididymal Secretory Proteins; D023083:beta-Defensins