Polyvinyl butyral (PVB) is a polymer resin byproduct of the automotive industry. Separation and recycling of PVB from automotive windshield production lines solve waste management issues and might be used in biomedicine implants, particularly when blended with high-density polyethylene (HDPE). Materials designed for biological applications must undergo pre-clinical safety evaluation, including in vitro biocompatibility and cytotoxicity testing. This study evaluated in vitro the biocompatibility and cytotoxicity of thick composite films made of recycled PVB (rPVB), HDPE, and composites made of concentrations of both polymers using peripheral blood mononuclear cells (PBMCs) from healthy donors. The microstructure and elemental analysis revealed no potential concerns regarding mechanical damage to the cell cultures. After culturing the PBMCs in the presence of the polymers and their blends, we observed no acute release of the proinflammatory cytokines TNF-α and IL1-β, nor evidence of cell death, measured by the release of nucleosomes to the extracellular medium at 24 h nor at 7 days of culture. We also found no damage to the cell monolayer. The stimulation with lipopolysaccharide (LPS), used here as proinflammatory control, induced the significant release of both cytokines and caused damage to the cell monolayer. In conclusion, the pure recycled PVB, pure HDPE, and their composites are safe and biocompatible.