Nanotechnology is an emerging technology which has wide applications in many fields including biomedical materials and antimicrobial applications. The study aimed to investigate the Antibacterial Activity of Synthesized Copper Nanoparticles from Caballero (Caesalpinia pulcherrima Swartz) Leaf Extract against gram positive bacteria Staphylococcus aureus and gram negative bacteria Escherichia coli. The antibacterial activity of copper nanoparticles from caballero leaf extract was assessed by determining its characteristics in terms of TEM Analysis and Morphological Examination which the average size of copper nanoparticles from caballero leaf extract to be 64.03 nm and all nanoparticle were spherical in shape. The antibacterial activity was tested using various concentrations (25%, revealed 50%, 75%, and 100%) of the copper nanoparticles from caballero leaf extract against gram negative bacteria Escherichia coli and gram positive bacteria Staphylococcus aureus. Clearing or Zone of inhibition was present in different concentration on the two types of bacteria. Antibacterial assay revealed that in different concentrations, higher or lower, of copper nanoparticles from caballero leaf extract has an antibacterial activity against gram negative bacteria Escherichia coli and gram positive bacteria Staphylococcus aureus. The difference in antibacterial activity of different concentration of copper nanoparticles from caballero leaf extract against gram negative bacteria Escherichia coli and gram positive bacteria Staphylococcus aureus using ANNOVA showed no significant difference. The study concluded that the synthesized Copper Nanoparticle from Caballero (Caesalpinia pulcherrima Swartz) Leaf Extract may be regarded as a potential antibacterial agent against both gram negative bacteria Escherichia coli and gram positive bacteria Staphylococcus aureus given at a higher or lower concentration.