Allergen immobilisation and signal amplification by quantum dots for use in a biosensor assay of IgE in serum

• Allergens immobilised by spotting on a polymer coated Dual Polarisation Interferometry (DPI) sensor chip.
• Activity and non-specific binding assessed over time.
• Detection of specific antibodies in serum demonstrated.
• Signal amplification using quantum dots pushes the limts of detection below 2.4 ng/mL of IgE in serum.

The production of biosensors for point of care diagnostics usually requires the immobilisation and storage of protein (for example, antigen or antibody) on a sensor surface, in a manner that retains a high degree of activity and low levels of non-specific binding. These characteristics have been assessed for polymer immobilised antigens (allergens) using an IgG binding assay and demonstrated further by assay with serum containing reactive IgEs.The activity of allergens immobilised on sensor chips using copoly(DMA–NAS–MAPS) and a spotting technique, as well as the specificity of their binding interactions with cognate immunoglobulins was assessed using Dual Polarisation Interferometry (DPI). The data obtained indicate that the allergens studied remain stable over long periods of time (at least 114 days). This performance compared favourably with other immobilisation methods. Allergen coated chips were tested in an anti-casein IgE assay using human serum from allergic and non-allergic donors. Detection of both total Ig and specific IgE was demonstrated using a secondary anti-IgE antibody. Furthermore, optical signal enhancement with streptavidin conjugated quantum dots was shown to yield responses for samples below 0.84 ng/mL (0.35 KU/L) of IgE, which overlap with the industrial quasi-standard ImmunoCAP® and is the clinically relevant threshold used to classify serum samples from allergic individuals.