7 alpha-Biotinylated testosterone derivatives as tracers for a competitive chemiluminescence immunoassay of testosterone in serum.
Ring core-biotinylated testosterone tracers were synthesized with bridges of three different lengths connecting the biotin moiety to the steroid core (7 alpha-Cn-Bio-T, n = 3, 6, or 11). Together with a position 7-specific polyclonal anti-testosterone antibody, we used the 7 alpha-C11-Bio-T tracer to develop a novel, labeled-hapten competitive immunoassay for total testosterone in serum. (The C3 and C6 tracers proved to be not suitable for analogous immunoassays.) Enhanced chemiluminescence signal was generated by use of a second immobilized antibody and a streptavidin-horseradish peroxidase conjugate. The measuring range of the assay is 0.2-20.0 nmol/L, linearity of serial dilutions can be demonstrated, the lower detection limit is 0.125 nmol/L, and the interassay imprecisions are 13-16%. Accuracy determinations in mass spectrometry-controlled reference specimens showed a mean recovery of 95%. In addition, the assay shows low cross-reactivities, demonstrating the favorable specificity of the combination of a "nearly native" tracer with a position analog antibody. The optimized steric structure and the long spacer arm of the biotinylated testosterone tracer make this chemiluminescence assay well-suited for measuring total testosterone concentration in serum.
Luppa
P
P
Institute of Clinical Chemistry and Pathobiochemistry, Klinikum rechts der Isar, Technical University Munich, Germany. peter.luppa@edv1.klinchem.med.tu-muenchen.de
Brückner
C
C
Schwab
I
I
Hauck
S
S
Schmidmayr
S
S
Birkmayer
C
C
Paulus
B
B
Hauptmann
H
H
D000293:Adolescent; D000328:Adult; D005260:Female; D006801:Humans; D007118:Immunoassay; D008163:Luminescent Measurements; D008297:Male; D008875:Middle Aged; D011863:Radioimmunoassay; D012680:Sensitivity and Specificity; D013739:Testosterone / Q000031:analogs & derivatives / Q000097:blood*
D013739:Testosterone