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Abstract: Poly(methacrylic acid-ethylene glycol dimethacrylate) was prepared using 2-aminopyridine as the imprinting molecule. This molecularly imprinted polymer (MIP) was ground and packed into a micro-column for selective solid phase extraction (SPE) of 2- aminopyridine from 20 mu l of sample solution. Non-specific adsorption was also confirmed for a structural analogue. Interestingly one of the isomers, 4-aminopyridine, bound most strongly to the MIP. The implication of resonance and basicity of this isomer molecule can be used to explain its strong binding with the self-assembled functional methacrylic acid (MAA) monomer. The monomer template complexion process was evaluated by Scatchard plots to determine a binding constant. The binding constant value is important for predicting the selectivity of a new MIP. After optimization of the molecular recognition process, a molecularly imprinted solid phase extraction-differential pulsed elution (MISPE-DPE) method was developed for the selective determination of 4-aminopyridine in serum with an analysis time of less than 3 min using a 2- aminopyridine micro-column for surrogate binding. The concentration detection limit was 0.5 μg/ml, which corresponded to an absolute detection limit of 10 ng. A larger sample volume of 845 μl afforded a better concentration detection limit of 52 ng/ml. (C) 2000 Elsevier Science B.V. All rights reserved