Ageratum conyzoides plays a role in traditional medicine in many parts of the world. It is widely use externally to treat skin diseases, wounds, ulcers, and boils, internally as febrifuge and to treat diarrhoe and haemorrhages. The reports related to bioassay of the extract of A. conyzoides is very limited. The objectives of the research is to study antibacteria activity from leaves and roots alkaloid A. conyzoides to growth of Staphylococcus aureus in vitro. Alkaloid leaves and roots extracted in methanol and dichloromethane as solvent. Crude extract analysed employing Gas Chromatography/Mass Spectroscopy (GC/MS). Antibacteria activity was studied by disc-diffusion and macro-dillution methods. The research used four concentrations, ie 30 g/ml, 40 g/ml, 50 g/ml, and 60 g/ml. Negative control used DMSO 1% and positive control used amphicillin 50 g/ml. GC/MS analysis showed that leaves extract consisted of 3, 8, 8-Trimethoxy-3-piperidyl-2, 2-binaphthyl-1, 14, 4-tetrone, Benzaldehide, 4-hydroxy-3, 5-dimethoxy,(4-hydroxy-3, 5-dimethoxyphenyl) methylene hyd), meanwhile roots extract consisted of 3, 3 Dimethyl-2-phenyl-2-(1-oxo-1, 2, 3, 4-Tetrahydronaphthalen-2-yl) Azirane). Leaves alkaloid of A. conyzoides revealed the highest inhibtion zone at 50 g/ml (10, 17±0, 24) mm, meanwhile roots alkaloid revealed the highest inhibition zone at 40 g/ml (10, 19±0, 33) mm. Minimum Inhibitory Cencentration (MIC) of leaves alkaloid is at 30 g/ml and MIC of roots alkaloid is at 25 g/ml. Minimum Bactericidal Concentration (MBC) of leaves alkaloid is at 40 g/ml, meanwhile MBC roots alkaloid is at 35 g/ml. Leaves and roots alkaloid statistically has the same effectively to inhibite growth of S. aureus. Leaves and roots alkaloid A. conyzoides could be used as traditional medicine for S. aureus infection.