A simple, rapid and cost effective preconcentration method is described for the determination of thallium in aqueous samples and human plasma using ion pair solvent microextraction (IP-SME) followed by flame atomic absorption spectrometry (FAAS). In trace element analysis, preconcentration and separation methods enhance the sensitivity and precision of the determination. In a preconcentartion step 10 ml of Tl solution was adjusted at pH=8 and treated with 2ml of 0.1 % 2-pyridyn mercaptan (PyM) transferred to a 15ml vial. Thallium formed a cationic complex with 2-pyridyn mercaptan. For creating ion pair and converted cationic complex to nonpolar form, 0.5 ml of 1% picric acid, as a counter ion, was added to the sample solution. The solution was stirred by a magnetic stirrer with a 6-mm bar at an optimized speed of 500 rpm. The ionpair, (Tl- PyM) + (picrate)- was formed and extracted by the acceptor phase consisting 4µl of nitrobenzene for 10 min. After extraction, the microdrop was retracted and to be measured by FAAS the drop volume was brought to 50 µl by acetone. Resulting solution directly injected into the nebulizer of AAS by a micro syringe for subsequent determination. Some effective parameters on extraction and complex formation, such as type and volume of organic solvent, pH, concentration of chelating agent and counter ion, extraction time and stirring rate were optimized. Under the optimum conditions, the enrichment factor and recovery were 188.0 and 94.0%, respectively. The calibration graph was linear in the range of 12 - 130 µg L-1 with correlation coefficient of 0.9989 under the optimum conditions of the recommended procedure. The detection limit based on the 3Sb criterion was 3.6 µg L-1 and relative standard deviation for (RSD) for ten replicate measurements of 50 µg L-1 and 100 µg L-1 lead was 4.9 and 4.7 % respectively. The results for determination of thallium in reference materials and human plasma demonstrated the accuracy, recovery and applicability of the presented method.