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. Author manuscript; available in PMC: 2010 May 1.
Published in final edited form as: J Immunol. 2009 May 1;182(9):5537–5546. doi: 10.4049/jimmunol.0803742

FIGURE 1.

FIGURE 1

Design of the Flk-1/VEGFR2-expressing Lm-based constructs. A, Cloned regions for each construct built are boxed; mapped CTL epitopes for the H-2d/q MHC I haplotype include the following: Flk210 (TYQSIMYIV), Flk613 (MFSNSTNDI), and Flk906 (PGGPLMVIV), and are shown in bold font (17). B, Each gene fragment was cloned into the expression vector pGG34 fused to LLO and placed under the control of the hly promoter. C, Western blot of proteins from culture supernatants showing expression of each fusion protein from the constructs listed. Polyclonal, rabbit, anti-PEST Ab was used for fusion protein detection (bottom), and mouse anti-LLO Ab was used for confirmation (top). Note that all lanes within a single blot were taken from the same Western blot; immunostaining was performed on two separate, but identical blots. D, IFN-γ ELISPOT showing CD8+ T cell-restricted responses ex vivo after immunization with each construct. The naive group was injected with PBS alone; all groups contained a control Lm group. Responses are to the corresponding mapped epitopes for each Flk fragment, as follows: pE1 = Flk-E1210–219 (TYQSIMYIV), pE2 = Flk-E2613–622 (MFSNSTNDI), and pI1 = Flk-I1906–915 (PGGPLMVIV); n = 5 per group. Graphs show mean ± SEM; *, p < 0.05; Mann-Whitney statistical test, experiment repeated once.