3. Detection of HLA -A2 restricted HER2 specific responses with MHC tetramers. 200 μl of whole blood is mixed with 10 μl HLA -A201 CMV pp65PE/anti -CD8 FITC (Beckman Coulter, San Diego, C A) or HER2 peptide specific MHC tetramer (Beckman Coulter, San Diego, CA) and incubated at room temperature for 30 minutes in the dark. 1 ml of OptiLyse C (Beckman Coulter) is added and incubated for 10 minutes to lyse the red cells. 1 ml of 1X PBS is ad ded and incubated for another 5 minutes. 2 mls 1X PBS is added and samples centrifuged. Cell pellet is resuspended in 500 μl 1% paraformaldehyde. The labeled cells are analyzed by flow cytometry and at least 500,000 events gated on lymphocytes are collected. C. Immunologic detection of antigen specific antibodies 1. Quantitative ELISA for the detection of HER2 protein sp ecific antibody responses . 96 well Immulon 4 plates (Baxter SP, Redmond, WA: Dynex Technologies) are incubated overnight at 4°C with a polyclonal affinity purified antibody to HER2 (kindly supplied by Dr. D.