A novel amperometric magnetoimmunosensor using an indirect competitive format is 
developed for the sensitive detection of the amino-terminal pro-B-type 
natriuretic peptide (NT-proBNP). The immunosensor design involves the covalent 
immobilization of the antigen onto carboxylic-modified magnetic beads (HOOC-MBs) 
activated with N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide (EDC) and 
N-hydroxysulfosuccinimide (sulfo-NHS), and further incubation in a mixture 
solution containing variable concentrations of the antigen and a fixed 
concentration of an HRP-labeled detection antibody. Accordingly, the target 
NT-proBNP in the sample and that immobilized on the MBs compete for binding to a 
fixed amount of the specific HRP-labeled secondary antibody. The 
immunoconjugate-bearing MBs are captured by a magnet placed under the surface of 
a disposable gold screen-printed electrode (Au/SPE). The amperometric responses 
measured at -0.10 V (vs. a Ag pseudo-reference electrode), upon addition of 
3,3',5,5'-tetramethylbenzidine (TMB) as electron transfer mediator and H2O2 as 
the enzyme substrate, are used to monitor the affinity reaction. The developed 
magnetoimmunosensor provides attractive analytical characteristics in 10-times 
diluted human serum samples, exhibiting a linear range of clinical usefulness 
(0.12-42.9 ng mL(-1)) and a detection limit of 0.02 ng mL(-1), which can be used 
in clinical diagnosis of chronic heart failure in the elderly and for 
classifying patients at risk of death after heart transplantation. The 
magnetoimmunosensor was successfully applied to the analysis of spiked human 
serum samples.