How to choose proper magnetic particles for bioaffinity interactions? The case for immobilised glyconanoconjugate

In this study, an assay for detection of the cancer biomarker Thomsen-nouvelle (Tn) antigen on the ELISA plates format was designed and developed. The effects of size and the interfacial density of the negative charge of magnetic beads (MBs) on the specific sensitivity of the bioaffinity interaction were studied. In particular, glyconanoconjugate, i.e. glycan Tn antigen conjugated to bovine serum albumin (BSA) was covalently immobilised on MBs for the bioaffinity detection of anti-Tn antibodies as cancer biomarkers. Six different MBs were used in the study, i.e. carboxy-modified MBs of 250 nm, 500 nm, 1000 nm and 2800 nm and epoxy-modified MBs of 2800 nm and 4500 nm. In order to evaluate which MBs are the best suited for detection of the analyte anti-Tn antibodies, sensitivities of detection (slopes from calibration curves) were calculated. Next, specific sensitivities were calculated for each type of MBs as a ratio of sensitivity of detection to the mass of MBs. From zeta potential ζ for each type of MBs, the interfacial charge density on MBs was calculated, expressed as the density of zeta potential ζ_d (ratio of zeta potential to surface area of MBs, i.e. ζ_d = ζ/A). Then, we evaluated the effect of size and ζ_d on the specific sensitivity of detection of anti-Tn antibodies in order to understand the immobilisation process on nanoscale. We also identified an optimal value of ζ_d on MBs; this was essential to achieve highly sensitive detection of the analyte, which made it possible to attain limit of detection (LOD) of (0.31 ± 0.01) ng mL^-1 or (2.10 ± 0.04) pM for analyte detection. In addition, the optimal assay configuration was highly selective and enabled reliable detection of the analyte in human serum with a recovery index in the range of 102-104%.