Comparison of CTL Memory and Recall Responses in Lung and Spleen after SQ or IN Prime-boost Vaccination with OVA.

<p>C57BL/6 mice were immunized intranasally with 10 μg of OVA in 50 μl PBS with 5% ADJ (SQ) or 10% ADJ (IN) twice at 3 week intervals. Lung and spleen were collected 3 weeks after the second vaccine. (<b>A-F)</b> Lung cells and splenocytes were stained with anti-CD8, anti-CD44, anti-CD62L, anti-CD103, anti-CD69, anti-CXCR3 and K^b/SIINFEKL tetramers; cells were acquired on a flow cytometer. (<b>A</b>) Absolute numbers of CD8 T cells in the lung and spleen. (<b>B</b>) Absolute numbers of SIINFEKL-specific tetramer-binding CD8 T cells in the lung and spleen. (<b>C</b>) CD103⁺ SIINFEKL-specific CD8 T cells in lung and spleen. (<b>D</b>) Absolute numbers of SIINFEKL-specific tetramer-binding resident memory-like CD103⁺CD69⁺CXCR3⁺ CD8 T cells in the lung and spleen. (<b>E-H</b>) Lung cells and splenocytes were stimulated with the SIINFEKL peptide and the percentages of IFN-γ⁺ IL-2⁺ TNF-α⁺ CD8 T cells were quantified by flow cytometry. (<b>E</b>) Frequency of SIINFEKL-specific IFN-γ⁺ CTLs among all CD8⁺ cells in lungs and spleen. (<b>F)</b> Absolute numbers of SIINFEKL-specific IFN-γ⁺ CTLs. (<b>G</b>) Frequency of SIINFEKL-specific IFN-γ⁺ IL-2⁺ TNF-α⁺ CTLs among all CD8⁺ cells. (<b>H)</b> Absolute numbers SIINFEKL-specific IFN-γ⁺ IL-2⁺ TNF-α⁺ CTLs. (<b>I-J)</b> C57BL/6 mice were immunized intranasally with 10 μg of OVA in 50 μl PBS with 5% ADJ (SQ) or 10% ADJ (IN) twice at 3 week intervals. 3 weeks after the second immunization, mice were challenged by intranasal administration of 500 PFU of recombinant influenza A/PR/8/34-OT-I H1N1 expressing the OVA SIINFEKL peptide. 6 days after challenge 5 mice/group were sacrificed and lungs were collected for quantification of CTLs and viral titration. (<b>I)</b> Frequency of SIINFEKL-specific IFN-γ⁺ CD8⁺ T cells among all CD8⁺ lung mononuclear cells. (<b>J)</b> Lung viral titers expressed as plaque-forming-units (PFU) per gram of lung. * indicates p<0.05, ** indicates p<0.01, *** indicates p<0.001. Data is representative of two experiments.