2-(Bis(pyridin-2-ylmethyl)amino) acetic acid (PIC) and ethylenediaminetetraacetic acid (EDTA) vehicle-based acetamidobenzoxazolone probes have been evaluated for the assessment of a 18 kDa translocator protein (TSPO). Here, PIC and EDTA work as a vehicle for possible binding of requisite metal and acetamidobenzoxazolone pharmacophore (MBP) has been used as a specific binding probe for TSPO. In silico interaction analysis was carried out to assess MBP's binding efficacy towards the TSPO. TSPO is a protein that is known to be overexpressed during inflammatory conditions, therefore, during in vivo experiments, an LPS-induced lung inflammation mouse model was developed for the administration of ^99mTc-PIC-MBP and ^99mTc-EDTA-MBP to understand absorption and distribution in several peripheral organs over time through SPECT imaging. In addition, photophysical analysis with serum protein, i.e., bovine serum albumin (BSA) was also performed using different spectroscopic techniques to understand the in vivo transportation/excretion of these two probes PIC-MPB and EDTA-MPB to better understand the nature of these probes as specific diagnostic agents.