This work demonstrated the preparation and characterization of a novel 4,4-difluoro-4-bora-3a,4a-diaza-s-indacene (BODIPY) derivative containing boronic acid group and investigation of its non-invasive/non-enzymatic fluorescence sensor behavior for determination of glucose. The novel BODIPY derivative bearing boronic acid pinacol ester (BODIPY-1) was synthesized by Sonogashira coupling reaction between Iodo-BODIPY and 4-ethynylphenylboronic acid pinacol ester. The target novel BODIPY-2 compound which was used as a fluorescence probe for the determination of glucose was synthesized from BODIPY-1 by changing the pinacol ester group to the boric acid moieties. The fluorescence intensity of the BODIPY-2 fluorophore decreased when it interacted with the glucose. Sensing performance towards to glucose of this probe was evaluated in detail concerning the suitable solvent, linear concentration range, convenient pH, limit of detection (LOD), limit of quantification (LOQ) and selectivity. The LOD value of BODIPY-2 was found 0.19 mM toward glucose. Also, the complex stoichiometry between the BODIPY-2 and glucose molecules was determined by Job’s plot technique.