Non-invasive imaging of living cells is an advanced technique that is widely used in the life sciences and medical research. We demonstrate a refractive index quantification microscopy (RIQM) that enables label-free studies of glioma cell-substrate contacts involving cell adhesion molecules and the extracellular matrix. This microscopy takes advantage of the smallest available spot created when an azimuthally polarized perfect optical vortex beam (POV) is tightly focused with a first-order spiral phase, which results in a relatively high imaging resolution among biosensors. A high refractive index (RI) resolution enables the RI distribution within neuronal cells to be monitored. The microscopy shows excellent capability for recognizing cellular structures and activities, demonstrating great potential in biological sensing and live-cell kinetic imaging.