Bacillus cereus was used as the assay organism for the quantification of chlortetracycline (CTC) in avian plasma. Antibiotic medium #8 gave significantly larger zones of inhibition than nutrient agar 1.5% when used as the assay medium (P less than or equal to 0.05). When the CTC concentration was measured in serum, citrated plasma, heparinized plasma, and oxalated plasma, no significant differences were found between the inhibition zone diameters produced on antibiotic medium #8. However, there was a significant decrease in the zone diameters produced on this medium when citrated whole blood, oxalated whole blood, and heparinized whole blood were used instead of plasma. The length of incubation of assay plates was inversely related to the inhibition zone diameter, with an incubation time of 8 hr giving the largest and most distinct zones. Storing prepared assay plates at 4 C for 24 hr before use appeared to decrease the variability of the inhibition zone diameters. Storage of citrated plasma at 4 C for 48 hr and at -60 C for 9 days resulted in no significant decreases in CTC concentrations.