Fluorescence polarization and anisotropy are two nearly equivalent techniques that have together, over the past 5 years, achieved wide use in high throughput screening in drug discovery. These are single-label methods that can be used to construct homogeneous assays that are fast, sensitive, and resistant to some significant interferences. Moreover, the assays are relatively inexpensive. This review surveys the peer-reviewed literature on the subject and explores some of the fundamental issues that bear on assay performance.
Sportsman, J.R., Lee, S.K., Dilley, H., Bukar, R. (1997). Fluorescence polarization in high throughput screening. In Devlin, J. (ed.), High Throughput Screening: the Discovery of Bioactive Substances, pp. 389-400. Marcel Dekker, New York.
3.
Pope, A.J., Haupts, U.M., Moore, K.J. (1999). Homogeneous fluorescence readouts for miniaturized high-throughput screening: Theory and practice. Drug Discov. Today4:350-362.
4.
Dandliker, W.B., DeSaussure, V.A. (1970). Fluorescence polarization in immunochemistry. Immunochemistry7:799-828.
5.
Lakowicz, J. (1999). Principles of Fluorescence Spectroscopy, 2nd ed.Kluwer Academic/Plenum, New York.
6.
Jolley, M. (1996). Fluorescence polarization assays for the detection of proteases and their inhibitors. J. Biomol. Screen.1:33-38.
7.
Kauvar, L.M., Higgins, D.L., Villar, H.O., Sportsman, J.R., Engqvist-Goldstein, A., Bukar, R., Bauer, K.E., Dilley, H., Rocke, D.M. (1995). Predicting ligand binding to proteins by affinity fingerprinting. Chem. Biol.2:107-118.
8.
Levine, L.M., Michener, M.L., Toth, M.V., Holwerda, B.C. (1997). Measurement of specific protease activity utilizing fluorescence polarization. Anal. Biochem.247:83-88.
9.
Lynch, B.A., Loiacono, K.A., Tiong, C.L., Adams, S.E., MacNeil, I.A. (1997). A fluorescence polarization based Src-SH2 binding assay. Anal. Biochem.247:77-82.
10.
Wu, P., Brasseur, M., Schindler, U. (1997). A high-throughput STAT binding assay using fluorescence polarization. Anal. Biochem.249:29-36.
Deshpande, S., Mineyev, I., Owicki, J. (1999). A robust, versatile tyrosine kinase assay for HTS in drug discovery. Prog. Biomed. Optics (SPIE)3603:251-261.
13.
Lynch, B., Minor, C., Loiacono, K., van Schravendijk, M., Ram, M., Sundaramoorthi, R., Adams, S., Phillips, T., Holt, D., Rickles, R., MacNeil, I. (1999). Simultaneous assay of Src SH3 and SH2 domain binding using different wavelength fluorescence polarization probes. Anal. Biochem.275:62-73.
14.
Zhao, G., Meier, T.I., Kahl, S.D., Gee, K.R., Blaszczak, L.C. (1999). BOCILLIN-FL, a sensitive and commercially available reagent for detection of penicillin-binding proteins. Antimicrob. Agents Chemother.43:1124-1128.
15.
Allen, M., Reeves, J., Mellor, G. (2000). High throughput fluorescence polarization: A homogeneous alternative to radioligand binding for cell surface receptors. J. Biomol. Screen.5:63-70.
16.
Banks, P., Gosselin, M., Prystay, L. (2000). Fluorescence polarization assays for high throughput screening of G protein-coupled receptors. J. Biomol. Screen.5:159-168.
17.
Banks, P., Gosselin, M., Prystay, L. (In press). Impact of a red-shifted dye label for high-throughput fluorescence polarization assays of G protein-coupled receptors. J. Biomol. Screen.
18.
Keating, S., Marsters, J., Beresini, M., Ladner, C., Zioncheck, K., Clark, K., Arellano, F., Bodary, S. (2000). Putting the pieces together: Contribution of fluorescence polarization assays to small molecule lead optimization. Prog. Biomed. Optics Imaging (SPIE)3913:128-137.
19.
Lee, P., Bevis, D. (In press). Development of a homogeneous high-throughput fluorescence polarization assay for G protein-coupled receptor binding. J. Biomol. Screen.
20.
Li, Z., Mehdi, S., Patel, I., Kawooya, J., Judkins, M., Zhang, W., Diener, K., Lozada, A., Dunnington, D. (2000). An ultra-high throughput screening approach for an adenine transferase using fluorescence polarization. J. Biomol. Screen. 5:31-37.
21.
Parker, G.J., Law, T.L., Lenoch, F.J., Bolger, R.E. (2000). Development of high throughput screening assays using fluorescence polarization: Nuclear receptor-ligand-binding and kinase/phosphatase assays. J. Biomol. Screen.5:77-88.
22.
Wu, J.J., Yarwood, D.R., Pham, Q., Sills, M.A. (2000). Identification of a high-affinity antiphosphoserine antibody for the development of a homogeneous fluorescence polarization assay of protein kinase C. J. Biomol. Screen. 5:23-30.
23.
Seethala, R., Menzel, R. (1998). A fluorescence polarization competition immunoassay for tyrosine kinases. Anal. Biochem.255:257-262.
24.
Zhang, J.-H., Chung, T., Oldenburg, K. (1999). A simple statistical parameter for use in evaluation and validation of high throughput screening assays. J. Biomol. Screen. 4:67-73.
25.
Bevington, P.R., Robinson, D.K. (1992). Data Reduction and Error Analysis for the Physical Sciences, 2nd ed.. chpt. 3. McGraw Hill, New York.
26.
Wei, A.-P., Herron, J.N. (1993). Use of synthetic peptides as tracer antigens in fluorescence polarization immunoassays of high molecular weight analytes. Anal. Chem.65:3372-3377.
27.
Terpetschnig, E., Szmacinski, H., Lakowicz, J.R. (1995). Fluorescence polarization immunoassay of a high-molecular-weight antigen based on a long-lifetime Ru-ligand complex. Anal. Biochem.227:140-147.
28.
Szmacinski, H., Castellano, F.N., Terpetschnig, E., Dattelbaum, J.D., Lakowicz, J.R., and Meyer, G.J. (1998). Long-lifetime Ru(II) complexes for the measurement of high molecular weight protein hydrodynamics. Biochim. Biophys. Acta1383:151-159.
29.
Kenakin, T. (1997). Pharmacologic Analysis of Drug-Receptor Interaction, 3rd ed., chpt. 10. Lippincott-Raven, Philadelphia.
30.
Bond, B., Ashman, S., Turconi, S., Brown, M., Hutchinson, J., Haupts, U., Rudiger, M., Bingham, R., Jurewsicz, T., Macarron, R., Earnshaw, D., Murray, K., Pope, A. (2000). Exploration of the relationships between test compound optical properties and pat teams of interference. Poster presented at the 6th Annual Conference of the Society for Biomolecular Screening. Vancouver. BCSeptember 6-9, 2000.
31.
Benson, R.C., Meyer, R.A., Zaruba, M.E., McKhann, G.M. (1979) Cellular autofluorescence is it due to flavins?J. Histochem. Cytochem.27:44-48.
32.
Aubin, J.E. (1979). Autofluorescence of viable cultured manmalian cells. J. Histochem. Cytochem. 27:36-43.
33.
van de Hulst, H. (1981). Light Scattering by Small Particles. chpt. 6. Dover, New York.
34.
Lakowicz, J.R., Gryczynski, I., Gryczynski, Z. (1999). High throughput screening with multiphoton excitation. J. Biomol. Screen.4:355-362.
35.
Moore, K.J., Surconi, S., Asdhman, S., Ruediger, M., Haupts, U., Emerick, V., Pope, A.J. (1999). Single molecule detection technologies in miniaturized high throughput screening: Fluorescence correlation spectroscopy. J. Biomol. Screen. 4:335-353.
36.
Weiss, S. (1999). Fluorescence spectroscopy of single biomolecules. Science283:1676-1683.
37.
Moerner, W.E., Oritt, M. (1999). Illuminating single molecules in condensed matter. Science283:1670-1676.
38.
Lakowicz, J.R., Gryczynski, I., Gryczynski, Z. (2000). Novel fluorescence sensing methods for high throughput screening. J. Biomol. Screen. 5:123-131.
39.
Coffin, J.R.Lataev, M., Bi, X., Nikiforov, T.T. (2000). Detection of phosphopeptides by fluorescence polarization in the presence of cationic polyamino acids: Application to kinase assaysAnal. Biochem.278:206-212.
40.
Perrin, F. (1926). Polarization de la lumiere de fluorescence: Vie moyenne de molecules dans letat excite. J. Phys. Raditim7:390.
41.
Rajkowski, K.M., Cittanova, N. (1981). Corrected equations for the calculation of protein-ligand binding results from fluorescence polarization data. J. Theor. Biol.93:691-696.