Substrate-Labeled Fluorescent Immunoassay

The hydrolysis of a nonfluorescent enzyme substrate to a fluorescent product is widely utilized to measure the activity of a large number of enzymes. Binding of enzyme substrates by antibodies often protects the enzymatically labile bond from hydrolysis. By the combination of these two formats, the substrate-labeled fluorescent immunoassay (SLFIA) was developed.  

---

Figure 6.2. Chemical structure of 8-[3-(7-/)-galactosylcoumarin-3-carboxyamido )propyl ] theophylline (B) used in the homogeneous substrate-labeled fluorescent immunoassay for theophylline (A). (Reprinted from Ref. 3, with permission from the American Association for Clinical Chemistry.)...

In the past ten years, numerous applications of fluorescence methods for monitoring homogeneous and heterogeneous immunoassays have been reported. Advances in the design of fluorescent labels have prompted the development of various fluorescent immunoassay schemes such as the substrate-labeled fluorescent immunoassay and the fluorescence excitation transfer immunoassay. As sophisticated fluorescence instrumentation for lifetime measurement became available, the phase-resolved and time-resolved fluorescent immunoassays have also developed. With the current emphasis on satellite and physician s office testing, future innovations in fluorescence immunoassay development will be expected to center on the simplification of assay protocol and the development of solid-state miniaturized fluorescence readers for on-site testing. 

Methods for the Detection of Antigens/Antibodies Equilibrium and kinetic inhibition assays based upon fluorescence polarization, 70, 3 fluorescence excitation transfer immunoassay (FETI), 70, 28 indirect quenching fluoroimmunoassay, 70, 60 the homogeneous substrate-labeled fluorescent immunoassay, 70, 79 fluorescence immunoassays using plane surface solid phases (FIAPS), 70, 87. 

Fig. 4. Schematic representation of the principle of substrate-labeled fluorescent immunoassay (tda, Ames). [Cited and modified from Fig. B-5, Miyai, K., in Miyai etal., eds. (M10).]...

HPLC = high-performance liquid chromatography. SLFIA = substrate-labelled fluorescent immunoassay. 

S28 Walter, B. (1981). A unitized solid phase analytical element for detection of therapeutic drugs in serum by homogeneous substrate-labeled fluorescent immunoassay. Clin. Chem. 27, 1086, Abstr. 311. 

Burd, J.F., Ellis, P.B., Greenquist, A.C., Li, T.M., Morris, D.L., Rupchock, P.A., Sommer, R.G., Tyhach, R.J., Walter, B. and Zipp, A.P. (1983). Measurement of theophylline with the substrate-labeled fluorescent immunoassay and apoenzyme reactivation immunoassay system in solution and in solid phase reagent strips. In Avrameas, S. et al. (Eds), Immunoenzymatic Techniques Proc. 2nd Int. Symp. Immunoenzymatic Techniques, Cannes, 16-18 March. Elsevier Science Publishers, Amsterdam, pp. 239-246. 

TM Li, JL Benovic, RT Buckler, JF Burd. Homogeneous substrate labeled fluorescent immunoassay for theophylline in serum. Clin Chem 27 22, 1981. 

SG Thompson, JF Burd. Substrate labeled fluorescent immunoassay for amikacin in human serum. Antimicrob. Agents Chemother 18 264, 1980. 

B Rollman, et al. Dibekacin assay in serum by automated fluorescence polarization immunoassay (Abbot TDX)—Comparison with high performance liquid chromatography, substrate labeled fluorescent immunoassay and radioimmunoassay. J Pharm Biomed Anal 4 53, 1986. 

A Comparison of Serum Phenytoin Determination by the Substrate-Labeled Fluorescence Immunoassay with Gas Chromatography, Liquid Chromatography, Radioimmunoassay and "EMIT ... 

---