This unit provides a detailed method for evaluating in situ DNA denaturation by flow cytometry. The principal technique is based upon the metachromatic properties of acridine orange. This technique has a number of advantages over traditional biochemical methods, but requires very precise methodology; the result is excellent clarity and differentiation between single-stranded and double-stranded DNA. This unit provides full details on the correct use of acridine orange together with an excellent discussion of the pitfalls and problems.