Performance evaluation of the phosphorescent porphyrin label: solid-phase immunoassay of alpha-fetoprotein.
Phosphorescent conjugates of antibodies, neutravidin, and biotin (pentylamine derivative) were synthesized using previously described monofunctional labeling reagent of platinum(II) coproporphyrin-I with isothiocyanate reactive group (PtCP-NCS). These conjugates, which can be considered as standard reagents for a range of bioanalytical applications, were evaluated in solid-phase immunoassay schemes with the clinical analyte a-fetoprotein (AFP). A custom-designed time-resolved phosphorescence plate reader based on a compact and low-cost 532-nm laser and optimized for measurement of porphyrin labels was used. Using optimized tracers, instrumentation and assay protocols, subpicomolar detection limits were obtained both for PtCP label in solution and for AFP in solid-phase immunoassay. This sensitivity is comparable with standard time-resolved fluorescence immunoassays with lanthanide labels. The performance of metalloporphyrin labels, instrumentation, and solid-phase immunoassays as an alternative to the established detection platforms is discussed.
O'Riordan
Tomás C
TC
Biochemistry Department, National University of Ireland, Cork, Lee Maltings, Prospect Row, Cork, Ireland.
Soini
Aleksi E
AE
Soini
Juhani T
JT
Papkovsky
Dmitri B
DB
D001710:Biotin / Q000737:chemistry; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D005470:Fluorometry; D006801:Humans; D007118:Immunoassay; D008163:Luminescent Measurements; D011166:Porphyrins / Q000737:chemistry; D015203:Reproducibility of Results; D000509:alpha-Fetoproteins / Q000032:analysis*
D011166:Porphyrins; D000509:alpha-Fetoproteins; D001710:Biotin