Detection of autoreactive CD4 T cells using major histocompatibility complex class II dextramers

The utility of MHC dextramers was evaluated in three autoimmune disease models: 1) proteolipid protein (PLP) 139-151-induced experimental autoimmune encephalomyelitis in SJL/J (H-2s) mice; 2) myelin oligodendrocyte glycoprotein (MOG) 35-55-induced experimental autoimmune encephalomyelitis in C57Bl/6 (H-2b) mice; and 3) cardiac myosin heavy chain (Myhc)-α 334-352-induced experimental autoimmune myocarditis in A/J (H-2a) mice. Flow cytometrically, we demonstrate that IAs/PLP 139-151, IAb/MOG 35-55 and IAk/Myhc-α 334-352 dextramers detect the antigen-sensitized cells with specificity, and with a detection sensitivity significantly higher than that achieved with conventional tetramers. Furthermore, we show that binding of dextramers, but not tetramers, is less dependent on the activation status of cells, permitting enumeration of antigen-specific cells ex vivo.The data suggest that MHC dextramers are useful tools to track the generation and functionalities of self-reactive CD4 cells in various experimental systems.Traditionally, limiting dilution analysis and cytokine ELISPOT assays have been used to enumerate the frequencies of antigen-specific cells, but low specificity and the tedious nature of these assays limit their use for routine applications [1-3]. To overcome these limitations, tetramer technology has been developed [4]. This approach involves derivation of fluorescent dye-labeled tetramerized complexes containing peptide-assembled major histocompatibility complex (MHC) molecules. The use of tetramers permits easy and rapid detection of antigen-sensitized T cells at a single cell level by flow cytometry (FC) [4,5]. Additionally, tetramer reagents can be used in conjunction with antibodies to phenotypically characterize the generation and expansion of antigen-specific cells during ensuing immune responses and to study their functionalities by cell sorting.Unlike with MHC class I tetramers, direct enumeration of antigen-specific cells - in particular, rare and l