Monitoring the immune response using real-time PCR.
Induction of an immune response to a particular antigen is the basis of vaccination. This has been done for years to prevent infectious diseases, and has the potential for the treatment of cancer. The immune response is nowadays more precisely modulated rather than simply induced, like in case of immunotherapy of allergic diseases. Likewise, autoimmune diseases are associated with an inappropriate immune response, and many efforts are made for specifically inhibiting this unwanted response. A possible line of attack is the induction of an antigen-specific immune tolerance, which also has a use in the field of transplantation, where allogeneic responses are deleterious for the graft. In all of these fields of fundamental and clinical medicine, the modulation of immune response requires the assistance of laboratory tests, among which real-time PCR appears more and more helpful. This chapter describes a protocol to quantify immune-related mRNAs using reverse transcription-real-time PCR. The transcripts can be quantified in cultured cells or in cultured whole blood, after an incubation period in the presence of the antigen to which the immune response is analyzed. This is the typical approach to evaluate the efficacy of a vaccine. The transcripts can also be quantified directly in the biological sample, giving information about the in vivo immune status of the individual. The techniques to achieve these different methods are described, and are illustrated by the analysis of the response against the toxoid tetanus antigen.
Stordeur
Patrick
P
Départment d'Immunobiologie-Hémobiologie-Transfusion, Hôpital Erasme, Université Libre de Bruxelles, Belgium.
D000818:Animals; D002478:Cells, Cultured; D020869:Gene Expression Profiling / Q000379:methods*; D005786:Gene Expression Regulation / Q000187:drug effects / Q000276:immunology*; D006801:Humans; D012333:RNA, Messenger / Q000096:biosynthesis / Q000276:immunology*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D013745:Tetanus Toxoid / Q000276:immunology* / Q000494:pharmacology; D014158:Transcription, Genetic / Q000187:drug effects / Q000276:immunology*
D012333:RNA, Messenger; D013745:Tetanus Toxoid