Doubly Phosphorylated Peptide Vaccines to Protect Transgenic P301S Mice against Alzheimer’s Disease Like Tau Aggregation
Abstract
:1. Introduction
2. Experimental
2.1. Solid Phase Peptide Synthesis
2.2. Synthesis of Phospho-Tau Peptide Vaccines
T cell epitope | Origin | Sequence * |
---|---|---|
TT | Tetanus toxin 582–599 of Clostridium tetani | VDDALINSTKIYSYFPSV |
TBC | Ag85B 241–255 of Mycobacterium tuberculosis | QDAYNAGGGHNAVFD |
B cell epitope | ||
Tau199–208[pS202/pT205] | hMAPT | SPGpSPGpTPGS |
Tau209–217[pT212/pS214] | hMAPT | RSRpTPpSLPT |
Tau229–237[pT231/pS235] | hMAPT | VRpTPPKpSPS |
2.3. Peptide Cleavage, Purification and Analysis
2.4. Animals
2.5. Immunization
2.6. Enzyme-Linked Immunosorbent Assay (ELISA)
2.7. Behavioral Characterization
2.7.1. Wire Hang Test
2.7.2. Accelerating Rotarod Test
2.7.3. Beam Walk Test
2.8. Primary Monoclonal Anti-phospho Tau Antibodies (mAb)
2.9. Immunohistochemistry
2.10. Preparation of Brain Homogenates
2.11. Relative Quantification of Phospho-Tau
2.12. Immunoblot Analysis
2.13. Statistical Data Analysis
3. Results
3.1. Design and Peptide Synthesis of Immunogens
B cell epitope | T cell epitope | tR [min] | MMcalc/MMobs | Yield [%] | Purity [%] |
---|---|---|---|---|---|
Tau199–208[pS202/pT205] | TT | 31.5 | 3368.6/3368.7 | 15.4 | 90 |
TBC | 15.3 | 2858.2/2858.1 | 13.8 | 90 | |
Tau209–217[pT212/pS214] | TT | 26.5 | 3539.8/3539.7 | 21.8 | 95 |
TBC | 18.8 | 3029.4/3029.4 | 22.8 | 85 | |
Tau229–237[pT231/pS235] | TT | 21.8 | 3493.8/3493.8 | 10.4 | >95 |
TBC | 26.9 | 2983.4/2983.6 | 9.8 | 95 |
3.2. Specific Total IgG Titers
3.3. Specific IgG1/IgG2a/IgG2b/IgG2c Titers
3.4. Tau Phosphorylation Degrees in Brain Homogenates
3.5. Immunohistochemical Quantification of Tau Pathology in P301S Mice
3.6. Behavioral Characterization of P301S Mice
4. Discussion
5. Conclusions
Supplementary Files
Supplementary File 1Acknowledgments
Author Contributions
Conflicts of Interest
References
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Richter, M.; Mewes, A.; Fritsch, M.; Krügel, U.; Hoffmann, R.; Singer, D. Doubly Phosphorylated Peptide Vaccines to Protect Transgenic P301S Mice against Alzheimer’s Disease Like Tau Aggregation. Vaccines 2014, 2, 601-623. https://doi.org/10.3390/vaccines2030601
Richter M, Mewes A, Fritsch M, Krügel U, Hoffmann R, Singer D. Doubly Phosphorylated Peptide Vaccines to Protect Transgenic P301S Mice against Alzheimer’s Disease Like Tau Aggregation. Vaccines. 2014; 2(3):601-623. https://doi.org/10.3390/vaccines2030601
Chicago/Turabian StyleRichter, Monique, Agneta Mewes, Manuela Fritsch, Ute Krügel, Ralf Hoffmann, and David Singer. 2014. "Doubly Phosphorylated Peptide Vaccines to Protect Transgenic P301S Mice against Alzheimer’s Disease Like Tau Aggregation" Vaccines 2, no. 3: 601-623. https://doi.org/10.3390/vaccines2030601