Identification of the minor H peptide HA-3. (A) CAD mass spectrum of candidate peptide (M+2H)²⁺ ion with monoisotopic m/z of 965.5 as eluted from Rp EBV-BLCL. (B) CAD mass spectrum of synthetic peptide VTEPGTAQY. Mass spectra were recorded on a Finnigan LCQ ion trap MS operating with a 3.0 atomic mass unit isolation window and 35% collision energy. The b and y ions are labeled above and below the amino acid sequence, respectively. Ions observed in the spectrum are underlined. (C) Minor H antigen HA-3 epitope reconstitution with synthetic peptides. A standard ⁵¹Cr-release assay was performed by incubating the indicated quantities of synthetic peptides with ⁵¹Cr-labeled HoDo target cells and then adding HA-3–specific CTLs. An E/T ratio of 17:1 was used. IVDCLTEMY corresponds to the minor H antigen A1-HY¹³ , and serves as a negative control. Background lysis of HoDo by the CTLs in the absence of any peptides was 7%; positive control lysis was 99%.