The present study offers a new method based on CdSe quantum dots (QDs) for simultaneous separation and determination of trace levels of heparin (Hep) in human serum samples. In this technique, CdSe QDs perform two different functions in Hep analysis process. Mercaptoacetic acid-capped red CdSe QDs
(λex=690 nm) are conjugated to Hep and the Hep-QD conjugation is then used as an extraction tool from microencapsulation of real samples. For quick release of the analyte from conjugation, the microcapsules are irradiated using an intensive near infrared wavelength as the controllable releasing agent. The Lcysteine-capped green CdSe QDs (λex=480 nm) play the additional role of Hep spectrofluorimetric detection. By labeling the appropriate chromophore, the extracted Hep can effectively quench green CdSe QD fluorescence which is quenched relative to Hep concentration. In comparison with other available methods, the newly-developed assay has a low detection limit (0.3 nmol L-1), wide linear range (1 to 8500 nmol L-1), good accuracy and high selectivity, that make it a good candidate for Hep separation and monitoring of complex physiological samples.