Figure 4
Chromatogram and spectra of TRITC-modified MIF chromatograms and spectra of the reference compound (TRITC bound to the N-terminal MIF peptide PMFIVNTNVPR) and fraction B from lymph node cell proteins isolated from TRITC-exposed mice and separated with 1D SDS-PAGE. The selected bands (see Fig. 3) were excised, digested and analyzed with UPLC-qToF. A reference compound was prepared to confirm the identity of the peptide. The qToF was operated in positive resolution mode with electrospray ionization. A qToF ms/ms method was used where the fixed mass was set to 865.9, the ms2 was set to 50–1500 m/z and the collision energy ramp was from 15 eV to 40 eV. The extracted chromatograms (a,b) correspond to the TRITC-fragment [m/z = 444.14, singly charged ion], the MIF peptide-fragment [m/z = 1287.68, singly charged ion] and the full TRITC-MIF conjugate [m/z = 865.91, doubly charged ion]. (a) Chromatogram of reference compound, (b) chromatogram of fraction B, (c) spectrum of reference compound and (d) spectrum of fraction B.
