Lymphocyte immunophenotyping

Specimen:

5-10 mL blood in lithium heparin, ACD or EDTA.

Method:

Peripheral blood mononuclear cells are incubated with fluorescent labelled monoclonal antibodies directed against specific lymphocyte surface antigens.

Detection of these antigens allows quantitation of total B cell numbers (CD19) and total T cell numbers (CD3), identification and quantitation of T cell subsets (CD4, CD8) and natural killer (NK) cells (CD16).

Cells are enumerated by Flow cytometry.

Reference Interval:

Affected by age and gender, diurnal rhythm and other physiological variables - consult pathologist.

See Table 5.

Application:

Assessment of immune deficiency states, lymphocytosis of unknown aetiology, diagnosis of lymphoid malignancies.

More extensive 'Flow cytometry' analysis is required for investigation of haematological malignancies.

Interpretation:

Lymphocyte numbers and CD4/8 ratio are interpreted in conjunction with clinical findings, lymphocyte morphology and other immune measurements.

In HIV infection, low absolute CD4 count (< 0.35 x 109/L) is predictive of disease progression, and < 0.20 x 109/L CD4 is associated with an increased risk of opportunistic infection.

T cells are increased in reactive disorders and in some infections (eg, Infectious mononucleosis).

B cells (CD19) are absent in X-linked hypogammaglobulinaemia but are usually present in common variable immunodeficiency. Increased B cell numbers occur in B cell proliferative diseases; monoclonality suggests leukaemia/lymphoma.

NK (CD16) cell numbers are reduced in some immunocompromised patients, patients with cancer, and in Chediak Higashi syndrome. NK cells may be increased in reactive conditions, hepatitis C virus infection, IV drug abuse, lymphomas.

See also Flow cytometry, Table 1, Table 2. 

Reference:

McCarthy DA et al. Cytometric Analysis of Cell Phenotype and Function. 2002. Cambridge University Press.