Introduction

In this chapter, some of the numerous assays for assessing cell function are described. Lymphocytes, monocytes, neutrophils, dendritic cells, and macrophages are important in the immune surveillance of microorganisms and malignant cells. All cell types are capable of secreting a variety of chemical mediators, and some are able to phagocytose particulate matter. Leukocyte recruitment to sites of infection is guided by complex phenomena that involve microbial emanations, cytokines, interleukins (ILs), chemokines, and microenvironment modifications of the vascular endothelium. The interaction between molecules on the surface of the leukocyte (CD11b and CD62L) with those on the endothelium (ICAM-1, CD62P, and CD62E) is critical for recruitment and attachment. After adherence to endothelium and prior to vascular emigration, monocytes and neutrophils undergo a rapid cytoskeleton rearrangement that is necessary for chemotaxis and for exerting their phagocytic properties against microorganisms. This latter step depends on the expression of specific receptors (FcRs, CD11b, CD 14, and CD35) that recognize opsonized (immunoglobulin-coated) microorganisms and particles.

From: Flow Cytometry: Principles and Applications Edited by: M. G. Macey © Humana Press Inc., Totowa, NJ

Micoorganisms and cellular material are phagocytosed into phagolysosomal vacuoles where they are subjected to strong oxygen-dependent micobicidal systems characterized by the so-called reduced nicotinamide adenine denucleotide (NADPH)-dependent respiratory burst and oxygen independent systems, including degrading enzymes, defensins (peptide antibiotics), and cathepsin G.

In addition to the their beneficial role, leukocytes (in particular, monocytes) may be involved in the pathogenesis of several noninfectious diseases such as acute and chronic myocardial ischemia, idiopathic pulmonary fibrosis, emphysema, rheumatoid arthritis, and certain forms of glomerulonephritis. The tissue-damaging properties arise when phagocyte activation mediated by upregulated membrane molecules results in degranulation with release of myeloperoxidase, elastase, and other proteases and by intermediate oxygen radicals.

One of the main obstacles to studying leukocyte function has been the difficulty encountered in (1) the transport of samples that require immediate analysis, (2) rapidly handling cells from peripheral blood and other sources such synovial fluid, bronchiolar lavage, and cerebrospinal fluid, (3) working with small volumes of blood or with blood from leukopenic individuals, (4) the complexity of some tests, and (5) processing several samples in a single work session. Many of these problems may be overcome by bringing the patient to the proximity of the laboratory and by using simple rapid assays to investigate the cells. Flow cytometry allows rapid assessment of the maturational stage of cells; expression of functional antigens, adhesion molecules, and receptors; priming; response to cytokines, chemoattractants, and activators, phagocytosis; and cell-cell interactions.

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