Mutations in the Ataxia Telangiectasia Mutated (ATM) gene are responsible for the autosomal recessive disease Ataxia Telangiectasia (A-T). A wide variety of mutations scattered across the entire coding region (9168bp) of ATM have been found, which presents a challenge in developing an efficient mutation screening strategy for detecting unknown mutations. Fluorescent chemical cleavage of mismatch (FCCM) is an ideal mutation screening method, offering a non-radioactive alternative to other techniques such as restriction endonuclease fingerprinting (REF). Using FCCM, we have developed an efficient, accurate and sensitive mutation detection method for screening RT-PCR products for ATM mutations. We have identified seven ATM mutations in five A-T families, four of which are previously unknown. We quantified ATM protein expression in four of the families and found variable ATM protein expression (0-6.4%), further evidence for mutant ATM protein expression in both classic and variant A-T patients. We conclude that FCCM offers a robust ATM mutation detection method and can be used to screen for ATM mutations in cancer-prone populations.