The article describes a method for the fast and effective purification of sequence-specific DNA binding proteins using DNA-agarose affinity chromatography. The method involves preparing chemically synthesized complementary oligodeoxynucleotides that contain a recognition site for the target protein, annealing and ligating these oligomers to form DNA oligomers. These oligomers are then covalently coupled to Sepharose CL-2B using cyanogen bromide to create an affinity resin. A partially purified protein fraction is combined with competitor DNA and passed through the DNA-Sepharose resin, allowing the desired protein to be purified due to its preferential binding to the recognition sites in the resin. The method has been successfully used to purify transcription factor Sp1, achieving a 500- to 1000-fold enrichment with an estimated 90% homogeneity and a 30% yield. The technique can also be applied to purify multiple DNA binding proteins simultaneously using tandem affinity columns with different binding sites. The authors discuss the importance of using appropriate competitor DNAs and buffer conditions for successful purification and outline a general strategy for the purification of sequence-specific DNA binding proteins.The article describes a method for the fast and effective purification of sequence-specific DNA binding proteins using DNA-agarose affinity chromatography. The method involves preparing chemically synthesized complementary oligodeoxynucleotides that contain a recognition site for the target protein, annealing and ligating these oligomers to form DNA oligomers. These oligomers are then covalently coupled to Sepharose CL-2B using cyanogen bromide to create an affinity resin. A partially purified protein fraction is combined with competitor DNA and passed through the DNA-Sepharose resin, allowing the desired protein to be purified due to its preferential binding to the recognition sites in the resin. The method has been successfully used to purify transcription factor Sp1, achieving a 500- to 1000-fold enrichment with an estimated 90% homogeneity and a 30% yield. The technique can also be applied to purify multiple DNA binding proteins simultaneously using tandem affinity columns with different binding sites. The authors discuss the importance of using appropriate competitor DNAs and buffer conditions for successful purification and outline a general strategy for the purification of sequence-specific DNA binding proteins.