8- to 12-week-old C57BL/6J mice (The Jackson Laboratory) were vaccinated with 50 μl of 25 mg/ml of AMCNPs or equivalent WCL vaccine subcutaneously via the hock on days 0, 2, and 4. On day 10, spleens were extracted and mechanically dissociated into single cell suspensions. Red blood cell lysis was performed by resuspending the cell mixture in ice cold ACK buffer (0.1 mM Na2EDTA, 10 mM KHCO3, 150 mM NH4Cl) for 5 min followed by washing with ice cold PBS and passage through a 40-μM cell strainer (Thermo Fisher Scientific). 5 × 106 splenocytes were then plated and cultured in 6-well suspension plates in BMDC growth media with 20 ng/ml GM-CSF and 1 μg/ml of either OVA SIINFEKL (InvivoGen) or WT1 RMFPNAPYL (MBL international, Woburn, MA) peptide. After 5 or 7 days of ex vivo culture, respectively, the supernatant was collected and analyzed for IFN-γ using ELISA kits (Biolegend, San Diego, CA). At 7 days of ex vivo culture, the splenocytes were stained as indicated (Supplementary Table 1 ) and used for flow cytometry analysis.
Ova siinfekl
Manufactured by InvivoGen
OVA SIINFEKL is a synthetic peptide fragment that contains the SIINFEKL sequence, a well-known CD8+ T cell epitope derived from the chicken ovalbumin protein. This peptide is commonly used in research applications as a model antigen for the study of CD8+ T cell responses.
Automatically generated - may contain errors
2 protocols using ova siinfekl
1
Evaluating T Cell Responses to Vaccine Formulations in Mice
2
Evaluating Vaccine-Induced Immune Responses
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8–12-week-old C57BL/6J mice (The Jackson Laboratory) were vaccinated with 50 μl of 25 mg/ml of AMCNPs or equivalent WCL vaccine subcutaneously via the hock on days 0, 2, and 4. On day 10, spleens were extracted and mechanically dissociated into single cell suspensions. Red blood cell lysis was performed by resuspending the cell mixture in ice cold ACK buffer (0.1 mM Na2EDTA, 10 mM KHCO3, 150 mM NH4Cl) for 5 min followed by washing with ice cold PBS and passage through a 40-μM cell strainer (Thermo Fisher Scientific). 5 × 106 splenocytes were then plated and cultured in six-well suspension plates in BMDC growth media with 20 ng/ml GM-CSF and 1 μg/ml of either OVA SIINFEKL (InvivoGen) or WT1 RMFPNAPYL (MBL international, Woburn, MA) peptide. After 5 or 7 days of ex vivo culture, respectively, the supernatant was collected and analyzed for interferon-γ (IFN-γ) using ELISA kits (Biolegend, San Diego, CA). At 7 days of ex vivo culture, the splenocytes were stained as indicated (Supplementary Table 1 ) and used for flow cytometry analysis.
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