WBC identification is usually done by Wright-stained peripheral blood smear examination. However, this approach creates problems due to the statistically small number of cells counted (usually 100), nonuniform cell distribution on the smear, and the need for subjective interpretation that can produce differences in cell counts in the same smear by the same technologist or between different technologists. Automated cell differential machines can improve the situation somewhat but currently still have problems with individual cells that are transitional between classification categories, atypical, or abnormal. In addition, neither a manual or machine differential can subtype normal or abnormal cells.

Flow Cytometry

Another approach to WBC counting is flow cytometry. Various WBC types and subtypes contain one or more antigens that are unique or are shared by a limited number of other cells. These antigens can be detected by specific monoclonal antibodies that can be tagged with a fluorescent molecule. A flow cytometer is able to activate the fluorescent molecule and detect, differentiate, and identify light wavelengths being produced. This permits detection, identification, and quantitation of the cells that possess the antigens being searched for. Usually an algorithmic approach is used in which one or two antibodies are tried, followed by one or two others depending on the initial results, and so on until final identification.

One problem (still present to some extent) was confusion because different manufacturers developed antibodies against the same or closely related cell antigens, but used different names for their antibody. Therefore, a standard nomenclature called cluster designation (CD) was developed in which each WBC antigen was given a CD number and the various antibodies (antibody “cluster”) that reacted with the same WBC antigen were assigned the corresponding CD number or numbers. That way, antibodies from various manufacturers, beside the proprietary brand name, could also be given a CD number that would indicate what antigen the antibody reacts with. Each antigen corresponds to a WBC category or subgroup. However, more than one CD antigen may be present on cells of the same WBC category or subgroup. For example, CD-4 antigen is found on the lymphocyte T-cell helper subgroup and CD-8 antigen on the lymphocyte T-cell suppressor subgroup. However, both CD-2 and CD-7 antigen are found on the lymphocyte T-cell natural killer subgroup. Certain platelet and megakaryocyte antigens are also included in the CD system.