False negative results resulting from antigen excess by immunophenotyping a leukemia patient with a whole blood...

A patient with prolymphocytic leukemia was immunophenotyped by whole blood and Ficoll-Hypaque methods. Discordance was noted between the two methods, especially for the HLE1, B1, I2, and T1B monoclonal markers. The disparity in results reflected the use of 15 X 10(6) cells versus 1 X 10(6) cells per reaction tube. This report represents a variable antigen excess problem with certain monoclonal antibodies while immunophenotyping a patient with leukemia by a whole blood flow cytometry method. The problem was resolved when both labeling methods used 1 X 10(6) cells per reaction tube. The results helped to encourage the vendor of the authors' commercial whole blood system to modify their procedure accordingly, but two other vendors have not yet made similar modifications. It suggests all commercial whole blood methods must be modified to accommodate for the leukocyte count and differential of the patient. Otherwise, the sensitivity of flow cytometers as used in traditional immunophenotyping protocols will not distinguish weakly stained cells from negative controls.