A cytochrome P450 3A4 (CYP3A4) based enzymatic biosensor was developed with the incorporation of a first-generation copper polypropyleneimine (CuPPI) metallodendrimer for the detection of anti-tuberculosis (anti-TB) drugs. The development of an electrochemical phenotype biosensor for this purpose is still vital since it aids in the ongoing fight against TB by determining metabolic profile. This allows TB treatment to be tailored on an individual patient basis, minimise adverse drug reactions and improve quality of life in TB patients. This simple biosensor was constructed via physical adsorption of CuPPI onto a gold electrode with subsequent electrostatic attachment of CYP3A4. The biosensor was successful in detecting all four first line anti-TB drugs i. e. isoniazid, ethambutol, pyrazinamide and rifampicin with limits of detection ranging from 0.02244 to 0.1072 nM in 0.1 M phosphate buffer. The developed biosensor was then applied towards "real samples" in the form of spiked synthetic urine and plasma. Calibration curves were carried out in the complex matrices, which were diluted with 0.1 M PB. These yielded good LOD in the range of ultra-low micromolar concentration i. e. 0.165-0.884 μM across all drugs. Recovery studies were also successful when detecting the real tablets in both plasma and urine with results ranging from 91.5 % to 108.5 %.