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Using RNA Aptamers and the Proximity Ligation Assay for the Detection of Cell Surface Antigens

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Biosensors and Biodetection

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 504))

Summary

The detection and typing of tumor cells based on differentially or similarly expressed antigens (biomarkers) have proven to be increasingly important for the diagnosis and treatment of various cancers. Sensitive techniques for the detection of cell surface antigens are therefore crucial for the early and accurate detection of cancer. Although techniques such as ELISA and tissue staining have proven their worth, these techniques often either require substantial amounts of starting material or are prone to high background and false negatives. The proximity ligation assay (PLA) has proven to be an exquisitely sensitive technique with very low background. Two probes that bind adjacent to one another on a protein target can be ligated, yielding a unique amplicon that can be sensitively detected by real-time PCR. We have now adapted PLA to cell surface protein targets using modified RNA aptamers, and have shown that aptamer-based cell surface PLA can successfully detect and differentiate between cells that differentially express a tumor antigen, the prostate specific membrane antigen (PSMA).

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References

  1. Chu, T., Ebright, J., and Ellington, A. D. (2007) Using aptamers to identify and enter cells. Curr Opin Mol Ther. 9, 137–144

    CAS  PubMed  Google Scholar 

  2. Jayasena, S. D. (1999) Aptamers: an emerging class of molecules that rival antibodies in diagnostics. Clin Chem. 45, 1628–1650

    CAS  PubMed  Google Scholar 

  3. Lee, J. F., Stovall, G. M., and Ellington, A. D. (2006) Aptamer therapeutics advance. Curr Opin Chem Biol. 10, 282–289

    Article  CAS  PubMed  Google Scholar 

  4. Nimjee, S. M., Rusconi, C. P., and Sullenger, B. A. (2005) Aptamers: an emerging class of therapeutics. Annu Rev Med. 56, 555–583 5. Yan, A. C., Bell, K. M., Breeden, M. M., and Ellington, A. D. (2005) Aptamers: prospects in therapeutics and biomedicine. Front Biosci. 10, 1802–1827

    Article  CAS  PubMed  Google Scholar 

  5. Yan, A. C., Bell, K. M., Breeden, M. M., and Ellington, A. D. (2005) Aptamers: prospects in therapeutics and biomedicine. Front Biosci. 10, 1802–1827

    Article  CAS  PubMed  Google Scholar 

  6. Chu, T. C., Marks, J. W. III, Lavery, L. A., Faulkner, S., Rosenblum, M. G., Ellington, A. D., and Levy, M. (2006) Aptamer:toxin conjugates that specifically target prostate tumor cells. Cancer Res. 66, 5989–5992

    Article  CAS  PubMed  Google Scholar 

  7. Chu, T. C., Twu, K. Y., Ellington, A. D., and Levy, M. (2006) Aptamer mediated siRNA delivery. Nucleic Acids Res. 34, e73

    Article  PubMed  Google Scholar 

  8. Daniels, D. A., Chen, H., Hicke, B. J., Swiderek, K. M., and Gold, L. (2003) A tenascin-c aptamer identified by tumor cell selex: Systematic evolution of ligands by exponential enrichment. Proc Natl Acad Sci U S A. 100, 15416–15421

    Article  CAS  PubMed  Google Scholar 

  9. Hicke, B. J., Marion, C., Chang, Y. F., Gould, T., Lynott, C. K., Parma, D., Schmidt, P. G., and Warren, S. (2001) Tenascin-c aptamers are generated using tumor cells and purified protein. J Biol Chem. 276, 48644–48654

    Article  CAS  PubMed  Google Scholar 

  10. Shangguan, D., Li, Y., Tang, Z., Cao, Z. C., Chen, H. W., Mallikaratchy, P., Sefah, K., Yang, C. J., Tan, W. (2006) Aptamers evolved from live cells as effective molecular probes for cancer study. Proc Natl Acad Sci U S A. 103, 11838–11843

    Article  CAS  PubMed  Google Scholar 

  11. Fredriksson, S., Gullberg, M., Jarvius, J., Ols-son, C., Pietras, K., Gustafsdottir, S. M., Ost-man, A., and Landegren, U. (2002) Protein detection using proximity-dependent DNA ligation assays. Nat Biotechnol. 20, 473–477

    Article  CAS  PubMed  Google Scholar 

  12. Gullberg, M., Fr edriksson, S., Taussig, M., Jar vius, J., Gustafsdottir, S., Landegren, U. (2003) A sense of closeness: protein detection by proximity ligation. Curr Opin Biotechnol. 14, 82–86

    Article  CAS  PubMed  Google Scholar 

  13. Gullberg, M., Gustafsdottir, S. M., Schall-meiner, E., Jarvius, J., Bjarnegard, M., Bet-sholtz, C., Landegren, U., and Fredriksson, S. (2004) Cytokine detection by antibody-based proximity ligation. Proc Natl Acad Sci U S A. 101, 8420–8424

    Article  CAS  PubMed  Google Scholar 

  14. Gustafsdottir, S. M., Schallmeiner, E., Fredriks-son, S., Gullberg, M., Soderberg, O., Jarvius, M., Jarvius, J., Howell, M., and Landegren, U. (2005) Proximity ligation assays for sensitive and specific protein analyses. Anal Biochem. 345, 2–9

    Article  CAS  PubMed  Google Scholar 

  15. Schallmeiner, E., Oksanen, E., Ericsson, O., Spangberg, L., Eriksson, S., Stenman, U. H., Pettersson, K., and Landegren, U. (2007) Sensitive protein detection via triple-binder proximity ligation assays. Nat Methods. 4, 135–137

    Article  CAS  PubMed  Google Scholar 

  16. Zhu, L., Koistinen, H., Wu, P., Narvanen, A., Schallmeiner, E., Fredriksson, S., Landegren, U., and Stenman, U. H. (2006) A sensitive proximity ligation assay for active PSA. Biol Chem. 387, 769–772

    Article  CAS  PubMed  Google Scholar 

  17. Pai, S., Ellington, A. D., and Levy, M. (2005) Proximity ligation assays with peptide conjugate ‘burrs’ for the sensitive detection of spores. Nucleic Acids Res. 33, e162

    Article  PubMed  Google Scholar 

  18. Lupold, S. E., Hicke, B. J., Lin, Y., and Cof-fey, D. S. (2002) Identification and characterization of nuclease-stabilized RNA molecules that bind human prostate cancer cells via the prostate-specific membrane antigen. Cancer Res. 62, 4029–4033

    CAS  PubMed  Google Scholar 

  19. Padilla, R., and Sousa, R. (1999) Efficient synthesis of nucleic acids heavily modified with non-canonical ribose 2′-groups using a mutant T7 RNA polymerase (RNAP). Nucleic Acids Res. 27, 1561–1563

    Article  CAS  PubMed  Google Scholar 

  20. Chu, T. C., Shieh, F., Lavery, L. A., Levy, M., Richards-Kortum, R., Korgel, B. A., and Ellington, A. D. (2006) Labeling tumor cells with fluorescent nanocrystal-aptamer bio-conjugates. Biosens Bioelectron. 21, 1859–1866

    Article  CAS  PubMed  Google Scholar 

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Pai, S.S., Ellington, A.D. (2008). Using RNA Aptamers and the Proximity Ligation Assay for the Detection of Cell Surface Antigens. In: Rasooly, A., Herold, K.E. (eds) Biosensors and Biodetection. Methods in Molecular Biology™, vol 504. Humana Press. https://doi.org/10.1007/978-1-60327-569-9_21

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  • DOI: https://doi.org/10.1007/978-1-60327-569-9_21

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-60327-568-2

  • Online ISBN: 978-1-60327-569-9

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