The alkaline phosphatase-streptavidin (AP-SA) probe released adenosine (∼267.2 Da) from the substrate adenosine monophosphate, where a signal may be detected from as little as 0.5 micro litres of a 0.
1 pg/ml dilution of the probe (2.6x10-22 mole). The signal from the AP-SA probe was linear from 1 to 50 pg/ml by monitoring adenosine release at 268 m/z (M+H) with liquid chromatography, electrospray ionization and quadrupole mass spectrometry (LC-ESI-MS). The safe limit of detection and quantification of the AP-SA probe was about 0.5 pg/well or 5 pg/mL. Enzyme linked Immuno mass spectrometric assay (ELIMSA) using the AP-SA probe provided a linear signal response for prostate specific antigen (PSA) against external standards from 1 to 500 pg/ml. The ELIMSA showed a safe limit of detection and quantification at 5 pg PSA per well or 50 pg/mL (false positive detection rate p≤0.01). Female samples of 100ul plasma per well were read against standards and blanks made in normal female plasma and the lowest sample quantified was about 9.8 pg/well or 98 pg/ml. Here ELIMSA was applied to measure PSA in plasma from female, normal male, prostatectomy patient and cancer patient samples that showed significant differences by ANOVA.
Analytical biochemistry. 2016 Mar 01 [Epub ahead of print]
Angelique Florentinus-Mefailoski, John G Marshall
Department of Chemistry and Biology, Ryerson University, 350 Victoria Street, Toronto, Canada., Department of Chemistry and Biology, Ryerson University, 350 Victoria Street, Toronto, Canada.