Immunoassay Using Fluorescent Dye-Trapped Liposomes Liposome Immune Lysis Assay (LILA)

  • Tatsuji Yasuda
  • Yoshio Ishimori
  • Mamoru Umeda


The liposome immune lysis assay (LILA) system was developed by S. C. Kinsky and coworkers in 1968 (Haxby, 1968). They used glucose as an internal liposome marker, and measured the glucose release in an enzymatic reaction. They also introduced umbelliferone phosphate and alkaline phosphatase as the first fluorescent marker system (Six, 1974). The enzyme reaction resulted in both the generation and the amplification of fluorescence. In 1977, Smolarsky et al. developed a novel LILA system using a complex of a fluorescent molecule (1-aminonaphthalene-3,6,8-trisulfonic acid (ANTS)) and a quencher (dipyridinium-p-xylene) as a marker trapped in liposomes. The complex, when entrapped in liposomes, shows little fluorescence. Upon lysis of the liposomes, dilution of the quencher in the external volume leads to a high fluorescence signal. One of the problems of the use of ANTS as a fluorophore is, however, that its excitation and emission wavelengths coincide with those of a fluorescent component existing in complement sources, and precise measurements may not be possible because of an increase in the background fluorescence.


Alternative Complement Pathway Sandwich Assay Complement Source Lipid Antigen Glycolipid Antigen 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Alving, C. R., Shichijo, S. and Mattsby-Baltzer, I. (1984) Preparation and use of liposomes in immunological studies. in “Liposome Technology”, G. Gregoriadis ed., CRC Press, Boca Raton, FL, 2: 157–175.Google Scholar
  2. Endo, T., Scott, D. D., Stewart, S., Kundou, S. K. and Marcus, D. M. (1984) Antibodies to glycosphingolipids in patients with multiple sclerosis and SLE. J. Immunol., 132: 1793–1797.PubMedGoogle Scholar
  3. Gerlach, E. and Deuticke, B. (1964) Eine einfache Methode zur Mikrobestimmung von Phosphat in der Papierchromatographie. Biochem. Z., 337: 477–479.Google Scholar
  4. Haxby, J. A., Kinsky, C. B. and Kinsky, S. C. (1968) Immune response of a liposomal model membrane. Proc. Natl. Acad. Sci. USA, 61: 300–307.PubMedCrossRefGoogle Scholar
  5. Ishimori, Y., Yasuda, T., Tsumita, T., Notsuki, M., Koyama, M. and Tadakuma, T. (1984) Liposome immune lysis assay (LILA): a simple method to measure Anti-protein antibody using protein antigen-bearing liposomes. J. Immunol. Methods, 75: 351–360.PubMedCrossRefGoogle Scholar
  6. Kabat, E.A. and Mayer, M.M. (1961) “Kabat and Mayer’s Experimental Immunochemistry”, Charles C. Thomas Pub., Springfield, IL, p. 133Google Scholar
  7. Kaise, S., Yasuda, T., Kasukawa, R., Nichimaki, T., Watarai, S. and Tsumita, T. (1985) Antiglycolipid antibodies in normal and pathogenic sera and synovial fluids. Vox Sang., 49: 292–300.PubMedCrossRefGoogle Scholar
  8. Kinsky, S. C. (1974) Preparation of liposomes and a spectrophotometric assay for release of trapped glucose marker. Methods in Enzymology, 32: 501–513.PubMedCrossRefGoogle Scholar
  9. Lelkes, P. I. (1984) Methodological aspects dealing with stability measurements of liposomes in vitro using the carboxyfluorescein. in “Liposome Technology”, G. Gregoriadis ed., CRC Press, Boca Raton, 3: 225–246.Google Scholar
  10. Leserman, L. D., Mache, P. and Barbet, J. (1981) Targeting to cells of fluorescent liposomes covalently coupled with monoclonal antibody or protein A. Nature, 288: 602–604.CrossRefGoogle Scholar
  11. Okada, N., Yasuda, T., Tsumita, T. and Okada, H. (1982) Activation of the alternative complement pathway of guinea-pig by liposomes incor porated with trinitrophenylated phosphatidylethanolamine. Immunology, 45: 115–124.PubMedGoogle Scholar
  12. Okada, N., Yasuda, T., Tsumita, T. and Okada, H. (1983a) Differing reactivity of human and guinea pig complement on haptenized liposomes via the alternative pathway. Mol. Inrounol., 20: 857–864.CrossRefGoogle Scholar
  13. Okada, N., Yasuda, T., Tsumita, T. and Okada, H. (1983b) Activation on the alternative complement pathway by natural antibody to glycollipids in guinea-pig serum. Immunology, 50: 75–84.PubMedGoogle Scholar
  14. Ralston, E., Hjelmeland, L. M., Klausner, R. D., Weinstein, J. N. and Blumenthal, R. (1981) Carboxyfluorescein as a probe for liposome-cell interactions: effect of impurities, and purification of the dye. Biochim. Biophys. Acta, 694: 133–137.Google Scholar
  15. Six, H. R., Young, W. W., Jr., Uemura, K. and Kinsky, S. C. (1974) Effect of antibody-complement on multiple vs. single compartment liposomes. Application of a fluorometric assay for following changes in liposomal permeability. Biochemistry, 13: 4050–4058.PubMedCrossRefGoogle Scholar
  16. Smolarsky, M., Teitelbaum, D., Sela, M. and Gitler, C. (1977) A simple fluorescent method to determine complement-mediated liposome immune lysis. J. Immunol. Methods, 15: 255–265.PubMedCrossRefGoogle Scholar
  17. Umeda, M., Ishimori, Y., Yoshikawa, K., Takada, M. and Yasuda, T. (1986a) Homogeneous determination of C-reactive protein in serum using liposome immune lysis assay (LILA). Jpn. J. Exp. Med., 56: 35–42.PubMedGoogle Scholar
  18. Umeda, M., Ishimori, Y., Yoshikawa, K., Takada, M. and Yasuda, T. (1986b) Liposome immune lysis assay (LILA): Application of sandwich method to determine a serum protein component with antibody bearing liposomes. J. Immunol. Methods, submitted.Google Scholar
  19. Weinstein, J. N., Yoshikami, S., Henkart, P., Blumenthal, R. and Higgins, W. A. (1977) Liposome-cell interaction: transfer and intracellular release of a trapped fluorescent marker. Science, 195: 489–492.PubMedCrossRefGoogle Scholar
  20. Weinstein, J. N., Ralston, E., Leserman, L. D., Klausner, R. D., Dragsten, P., Henkart, P. and Blumenthal, R. (1984) Self-quenching of carboxyfluorescein fluorescence: uses in studying liposomes stability and liposome-cell interaction. in “Liposome Technology”, G. Gregoriadis ed., CRC Press, Boca Ralton, FL, 3: 183–204.Google Scholar
  21. Yasuda, T., Naito, Y., Tsumita, T. and Tadakuma, T. (1981) A simple method to measure anti-glycolipid antibody by using complement-mediated immune lysis of fluorescent dye-trapped liposomes. J. Immunol. Methods, 44: 153–158.PubMedCrossRefGoogle Scholar

Copyright information

© Plenum Press, New York 1988

Authors and Affiliations

  • Tatsuji Yasuda
    • 1
  • Yoshio Ishimori
    • 1
  • Mamoru Umeda
    • 1
  1. 1.Laboratory of Biological Products, The Institute of Medical ScienceThe University of TokyoMinato-ku, Tokyo 108Japan

Personalised recommendations