Here we show that this inherent limitation can be solved using a relatively simple approach. This approach involves the development of a self-referenced biosensor consisting of two almost identically sized dye-doped polystyrene microspheres placed on adjacent holes at the tip of a suspended core optical fiber. Here self-referenced biosensing is demonstrated with the detection of Neutravidin in undiluted human serum samples. The fiber allows remote excitation and collection of the WGMs of the microspheres in a dip sensing setting. By taking advantage of surface functionalization techniques, one microsphere acts as a dynamic reference, compensating for nonspecific binding events, while the other microsphere is functionalized to detect the specific interaction. The almost identical size allows the two spheres to have virtually identical refractive index sensitivity and surface area. This ensures their responses to nonspecific binding and environmental changes are almost identical, whereby any specific changes such as binding events, can be monitored via the relative movement between the two sets of WGM peaks.
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