Paper in Analyst: Statistical approach for reproducible, sensitive and specific end-point detection of immunoreactions

In the biomedical field, end-point detection bioassays such as enzyme-linked immunosorbent assays (ELISAs) are essential tools because of their simplicity, high-throughput, and suitability for their use at the point-of-care. End-point bioassays are significantly constrained by the need of sample labeling with fluorescent or colorimetric tags for subsequent detection. A promising strategy to overcome these limitations is to harness recent advances in label-free biological nanosensors. Here we analyse the potential of nanomechanical biosensors based on surface stress for the end-point detection of horseradish peroxidase. Our study comprises the analysis of 1012 cantilevers with the technology SCALA (Mecwins S.L.). We find that the surface stress induced by the antibody-antigen binding exhibits is significantly correlated to the surface stress generated during the antibody attachment and blocking steps. The statistical correlation is harnessed to identify immobilization failure or success, and thus enhancing the specificity and sensitivity of the assay. This procedure enables achieving a rate of true positives and true negatives of 90% and 91% respectively. The detection limit is of 10 ng/mL (250 pM) that is similar to detection limit obtained in our ELISA and at least two orders of magnitude smaller than that achieved with well-established label-free biosensors such as quartz crystal microbalance (QCM) and surface plasmon resonance (SPR).


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