title: Considerations for the control of background fluorescence in clinical flow cytometry.
abstract: Accurate measurement of antigen-positive cells by flow cytometry can be hampered by background fluorescence of antigen-negative cells and other particles (e.g., debris). This article focuses on three major causes of background (autofluorescence, spectral overlap, and undesirable antibody binding) by reviewing individual aspects of flow cytometric measurements that contribute to these causes. The appropriate use of controls facilitates a thorough understanding of these contributing factors as well as the development of robust cell labeling protocols intended for routine flow cytometric analysis. We present a set of recommendations that enables the user to develop an optimized cell labeling protocol that minimizes background and maximizes the ability to reliably distinguish between a positive and a negative population of cells. These recommendations are also intended to augment existing guidelines designed to aid in the formulation of a consensus regarding the utility of flow cytometry for the analysis of clinical samples.
authors: Cytometry, Cytonome/ST, LLC, Boston, Massachusetts 02210, USA. rhulspas@cytonomest.com
authors: Ruud
authors: R
authors: Hulspas
authors: Maurice R G
authors: MR
authors: O'Gorman
authors: Brent L
authors: BL
authors: Wood
authors: Jan W
authors: JW
authors: Gratama
authors: D Robert
authors: DR
authors: Sutherland
mesh_terms: D000906:Antibodies / Q000378:metabolism; D000918:Antibody Specificity; D019411:Clinical Laboratory Techniques / Q000379:methods*; D005434:Flow Cytometry / Q000379:methods*; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D007091:Image Processing, Computer-Assisted; D011786:Quality Control; D013194:Staining and Labeling / Q000379:methods
chemical_list: D000906:Antibodies; D005456:Fluorescent Dyes