We have developed a novel method for genotyping based on free solution affinity capillary electrophoresis. We prepared DNA-nanoparticle conjugates by mixing biotin-modified DNA and NeutrAvidin-modified polystyrene nanoparticles; this mixture was then injected into a capillary. Subsequently, we injected the fluorescent-labeled sample DNAs into the capillary, applied the voltage, increased its temperature after 7 min, and detected the fluorescence at its anodic end. This novel method was applied for genotyping human c-K-ras, and the three genotypes were definitely distinguishable with high reproducibility. This method can be easily automated, and it is useful for high-throughput gene mutation analysis.