Using nano-size glass flakes as a model and Poly (vinyl alcohol) (PVA) as polymer matrix, glass flake-based antibacterial nanocomposite materials were successfully developed through layer-by-layer method. Characterization by ash test revealed a high percentage of nanofiller content which is in the range from 70wt. % to 79 wt. %. Scanning electron microscopy (SEM) results indicated the successful preparation of the multilayer thin films with thickness from 8 um to 65 um. In addition, antibacterial test results revealed that the glass flake-based antibacterial nanocomposite materials possessed very good antibacterial properties against both gram-negative bacteria E. coli and gram-positive bacteria S. aureus with the killing percentage of 96.5% and 96.9% at the concentration of 5% biocide. Therefore, it could be a promising antibacterial insulating paint for power grid applications.