Performance of label-free biosensors as a function of layer thickness

Biosensors are used in various applications in the field of medicine, environmental monitoring, and chemical processing to study the concentration and quality of target molecules. Biosensors convert the interaction between a specific target molecule and a recognition element into a measurable signal. We compared the performance of four different label-free biosensor techniques (multi-parametric surface plasmon resonance (MP-SPR), quartz crystal microbalance (QCM), mass-sensitive micro array (MSMA, or also known as film bulk acoustic resonators (FBARs)) and biolayer interferometry (BLI)) to evaluate how well they can quantify thick protein layers. We utilized the avidin-biotin system, which enables tight and specific binding and allowed us to assemble layers of proteins on the biosensor surface in a well-defined and reproducible fashion. Our results show that MP-SPR outperforms the other label-free biosensors in analyzing thick samples, showing a predictable and sensitive binding signal for over 50 albumin – avidin layers, which is estimated to correspond to a 300–400 nm thick protein layer. The linear measurement range of BLI was 38 layers corresponding to a 228–304 nm thick surface on the biosensor while QCM and MSMA were able to measure 108–144 nm and 72–96 nm thick protein layers with a fairly linear response, respectively. • Thick samples may cause troubles in biosensor measurements. • Comparison of biosensors with thick samples favors SPR technique. • The measured response does not directly reflect the layer thickness. • Feasibility of the biosensor technique is dependent on the aim of the assay.