DAPI, 4′,6-diamidino-2-phenylindole, binds to double-stranded DNA forming a complex that fluoresces up to twenty times more than DAPI alone. It is widely used in biochemical and cytochemical studies for DNA detection. The experiment described here measures the DAPI binding constant, Kƒ , for calf-thymus DNA using fluorescence measurements and a modified Scatchard analysis. This experiment also allows the student to estimate the occupancy of the minor groove by calculating the number of nucleotides per bound ligand. Modern pharmaceutical studies involve the generation of fluorescence titration curves by the addition of aliquots of DNA to a fixed quantity of the therapeutic agent or ligand. A Scatchard plot of the binding data is then calculated by plotting the ratio of bound/free ligand versus the bound fraction and calculating the slope. This experiment utilizes the same one-cuvette-type procedure, but employs a modified Scatchard analysis that has the student calculate a fraction, ƒ, that represents the fraction of bound ligand varying from zero to one. The modified Scatchard plot involves a reciprocal plot of DNA/ƒ versus 1/(1 − ƒ ), where the slope yields the binding constant Kƒ and the intercept gives a value for the number of nucleotides per ligand binding site.
Experiments, laboratory exercises, lecture demonstrations, and other descriptions of the use of chemicals, apparatus, instruments, computers, and computer interfaces are presented in the Journal of Chemical Education as illustrative of new or improved ideas or concepts in chemistry instruction and are directed at qualified teachers. Although every effort is made to assure and encourage safe practices and safe use of chemicals, the Journal of Chemical Education cannot assume responsibility for uses made of its published materials. Many chemicals are hazardous. Precautions for the safe use of hazardous chemicals and directions for their proper disposal are described in the Material Safety Data Sheets and on the labels. We strongly urge all those planning to use materials from our pages to make choices and to develop procedures for laboratory and classroom safety in accordance with local needs and situations.
Our Secondary School editors work hard to distill all the JCE materials to produce a fraction of particular interest to high school teachers. We call it CLIC.
In recent years we have worked hard to better match our advertisers with our readers. When shopping for chemistry education materials, visit our advertisers' WWW sites first.
Take JCE along on your outreach missions. Copies of the Journal, guest access to JCE Online, our publications catalog, and more are available for your participants.
Caution! 
