The aim of the present study was to develop a time-efficient chromatographic method for the analysis of therapeutic concentrations of ganciclovir (GCV) in plasma, urine as well as dialysate (from continuous renal replacement therapy) from solid organ transplant recipient treated with either GCV or its prodrug valganciclovir (VGCV) in combination with a wide variety of other concomitant drugs. Sample preparation was performed by reversed phase solid phase extraction and was followed by separation of the analytes on a reversed phase column using isocratic elution with a mobile phase consisting of acetonitrile—a counter ion (50 mM 1-heptanesulfonic acid) in an aqueous sodium dihydrogen phosphate buffer (pH 2.1; 10 mM) (10:90 v/v) and a fluorescence detector. Validation of the method showed linearity within the concentration range of 0.1–40 μg/mL for plasma and 0.1–120 μg/mL for urine and dialysate (R2 > 0.99, n ≥ 5). Accuracy and precision (evaluated at 0.1, 5 and 40 μg/mL) were both satisfactory. The LLOQ was determined to be 0.1 μg/mL. The method was successfully applied on clinical samples from renal transplant recipients treated with VGCV in combination with a variety of usually used concomitant drugs for solid organ transplant recipients.