A high performance thin-layer chromatographic (HPTLC) method for quantitative analysis of trazodone in human serum has been developed and validated. Trazodone was extracted with n-hexane: isoamyl alcohol (99:1). Chromatographic separation was performed on silica gel 60 F 254 HPTLC plates, with toluene: acetone: ethanol: ammonium (9:7:2:0.5, v/v/v/v) as mobile phase. Densitometric evaluation was carried out at 290 nm. There was no chromatographic interference between trazodone and its major metabolite, m-chlorophenilpiperazine (R f 0.82 and 0.39, respectively). Regression analysis of the calibration plot revealed good linearity (r=0.999) over the range of 20.00 and 200.00 ng/band, corresponding to 0.20 and 2.00 ng/μL of trazodone in human serum after extraction process and applying 10 μL to the chromatographic plates. The % RSD of intra-assay and inter-assay precision, were in the range of 0.98% to 2.97% (n=3) and 1.06% to 3.54% (n=9), respectively. LOD and LOQ were found to be 0.016ng/μL, and 0.048ng/μL, and the recovery values, of trazodone from serum, were between 92.52 % and 96.73 %. This method was successfully applied to quantify trazodone in patient serum samples. In conclusion, the developed method might be useful for the quantitative determination of trazodone in human serum, as a tool to evaluate patient adherence to their pharmacotherapy with this drug.