Dr. Enrico Lugli, cytometry unit manager and principal investigator of the laboratory of translational immunology at humanitas.

An important factor that must be considered for proper panel design is the sensitivity of the reagent. Sensitivity is influenced by the brightness of the fluorochrome and the data spread due to the presence of other antibody conjugates attached to the same cell. Not all fluorochromes have the same sensitivity due to differing brightness. Sensitivity is also impacted by the expression level of the markers being studied. Markers, or antigens, can be classified as primary, secondary or tertiary, depending on their levels of expression. Many candidate antibody-conjugate panels may need to be evaluated to determine which one optimally stains all the antigens under study. This is also important to determine which antibody conjugates reduce the sensitivity. Single stain controls to evaluate and correct for compensation must also be performed. Multicolor experiments can be frustrating but using the right panel can give wonderful results and it is important to invest time and effort into this.

Mahnke, Y.D,  Roederer, M, Clin. Lab. Med; 27(3): 469, (2007)