Light-analyte interaction systems are key elements of novel near-field optics based sensing techniques used for highly-sensitive detection of various kinds of targets. However, it is still a great challenge to achieve quantitative analysis of the targets using these sensing techniques, since critical difficulties exist on how to efficiently and precisely introduce the analytes into the desired location of the near-field light focusing, and quantitatively measure the enhanced optical signal reliably. In this work, we present for the first time a localized photonic nanojet (L-PNJ) based sensing platform which provides a strategy to achieve quantitative biosensing via utilizing a unique light-analyte interaction system. We demonstrate that individual fluorescent microsphere of different sizes can be readily introduced to the light-analyte interaction system with loading efficiency more than 70%, and generates reproducible enhanced fluorescence signals with standard deviation less than 7.5%. We employ this sensing platform for fluorescent-bead-based biotin concentration analysis, achieving the improvement on the detection sensitivity and limit of detection, opening the door for highly sensitive and quantitative biosensing. This L-PNJ based sensing platform is promising for development of next-generation on-chip signal amplification and quantitative detection systems.