The determination of plasma CETP activity with isotopically labeled lipoproteins is known to be laborious. We report a method to assess CETP activity in a defined system by using fluorescent cholesteryl linoleate analog in comparison to an isotopic assay.Method: Exogenous donor (MW 756) and acceptor particles (human VLDL) are combined with CETP source (plasma) within a total volume of a 10 mM tris buffer (pH 7.4). Donor particles consist of a nitrobenzoxadiol-fluorophor which takes advantage of a self-quenching effect on microemulsion incorporated cholesteryl linoleate (WAK, Germany). CETP mediated transfer is determined by the increase of fluorescence intensity as the cholesteryl linoleate is removed from the self quenched donor to the acceptor particle (incubation time 3 h at 37° C; EX 435 nm, EM 535 using a LS50 fluorescence photometer (Perkin Elmer)). A standard curve to derive the relation between fluorescence intensity and mass transfer is generated from a linear dilution of a defined concentration of the donor particle in isopropanol.Results: After screening of 50 plasma samples, a nearly 10 fold individual transfer rate between the lowest (6 pmol/3 h) and the highest (57 pmol/3 h) one was observed. 10 series of 10 samples and a series of 20 measures of 3 samples in different CET activities (15/34/46 pmol/3 h) show an interassay CV of 14.6%, and an intraassay CV of 10.8% (all samples were measured in duplicates). A good correlation is found between CET rates of 20 samples obtained from the fluorescent assay (range 5-65 pmol/3 h) and from the isotopic assay (r = 0.9, p = 0.000).Conclusion: Taken together, this new fluorimetric assay for determining CETP activity has the advantage of avoiding radioactive substrate. In addition, it provides a fast, simple and specific method for quantifying CETP activity in the clinical laboratory.