Enhanced sandwich immunoassay using antibody-functionalized magnetic iron-oxide nanoparticles for extraction and detection of soluble transferrin receptor on a photonic crystal biosensor

Iron deficiency anemia (IDA) has detrimental effects on individuals and societies worldwide. A standard sandwich assay (SA) for the detection of soluble transferrin receptor (sTfR), a biomarker of IDA, on a photonic crystal (PC) biosensor was established, but it was susceptible to non-specific signals from complex matrixes. In this study, iron-oxide nanoparticles (fAb-IONs) were used as magnetic immunoprobes to bind sTfR and minimize non-specific signals, while enhancing detection on the PC biosensor. This inverse sandwich assay (IA) method completely bound sTfR with low variability (< 4% RSD) in buffer and allowed for its accurate and precise detection in sera (Liquichek (TM) control sera) on the PC biosensor using two certified ELISAs as reference methods. A linear dose-response curve was elicited at the fAb-IONs concentration in which the theoretical binding ratio (sTfR:fAb-IONs) was calculated to be <1 on the IA. The LoDs for sTfR in the SA and IA were similar (P> 0.05) at 14 and 21 mu g/mL, respectively. The inherent imprecision of the IA and reference ELISAs was sigma(delta)=0.45 mu g/mL and the mean biases for Liquichek (TM) 1, 2 and 3 were 0.18, 0.19 and -0.04 mu g/mL, respectively. Whereas the inherent imprecision of the SA and reference ELISAs was sigma(delta)=0.52 mu g/mL and the biases for Liquichek (TM) 1, 2 and 3 were 0.66, 0.14 and -0.67 mu g/mL, respectively. Thus, unlike the SA, the IA method measures sTfR with the same bias as the reference ELISAs. Combined magnetic separation and detection of nutrition biomarkers on PC biosensors represents a facile method for their accurate and reliable quantification in complex matrixes. (C) 2015 Elsevier B.V. All rights reserved.