Figure 3

Intensity comparison between Gag CM9 Qdot 655 multimer and Gag CM9 APC-tetramer. A) Single cell suspension cells of a lymph node from an SIV infected Mamu-A*01 positive RM were stained with Gag CM9 Qdot 655 multimer or with Gag CM9 APC tetramer and the staining intensity was measured. Z stack average intensity projections for six cells was used and the mean average intensity was calculated by using Image J Software. A ten-fold higher mean average intensity was found for Gag CM9 Qdot 655 multimer as compared to the Gag CM9 APC tetramer, even though the same monomers are used in each case. B-D) Fluorescence images of tissue sections from an SIV infected Mamu-A*01 positive RM stained with CD8 in green, Gag CM9 Qdot 655 multimer in red (left column) or Gag CM9 APC Tetramer in red (right column) and dapi in blue. B) Images of spleen sections stained with Gag CM9 Qdot 655 multimer demonstrated abundant Gag CM9 specific cells (left column) whereas the consecutive section stained with Gag CM9 APC Tetramer showed no Gag CM9 positive cells (right column). C) Gag CM9 positive cells were detected in a solitary lymphoid follicle within the colon section (right column) when Gag CM9 Qdot 655 multimer was used, whereas when the consecutive slide was stained with Gag CM9 APC Tetramer no Gag CM9 positive cells could be detected (left column). D) Gag CM9 positive cells were detected both in the lamina propria and in a small solitary lymph node in the colon (right column) when stained with Gag CM9 Qdot 655 multimer but when the consecutive slide was stained with Gag CM9 APC tetramer no Gag CM9 positive cells were detected (left column). Images were collected with a 20×/0.75 objective. Scale bar = 50 μm.