CGG repeat expansions in the 5' noncoding region of the fragile X mental retardation 1 gene (FMR1) give rise to both neurodevelopmental and neurodegenerative human diseases depending on the length of the expansion. Expansions beyond 200 repeats (full mutation) generally result in gene silencing and fragile X syndrome (FXS), the leading heritable form of cognitive impairment and autism. Smaller expansions (55-200 CGG repeats; "premutation") give rise to the neurodegenerative disorder fragile X-associated tremor/ataxia syndrome (FXTAS) through an entirely distinct, toxic mRNA gain-of-function mechanism. A rapid means for both high-risk and newborn screening for allele size would provide a greater opportunity for early intervention and family counseling as well as furnish critical data on repeat size distribution and expanded allele frequencies. In the current work, we propose a novel mass spectrometry (MS) based method for the rapid identification of expanded CGG repeats to complement a recently described polymerase chain reaction (PCR) method for large population screening. In this combined approach, the optimized PCR method is used to amplify the relevant region of FMR1, followed by extensive nonspecific nuclease digestion. The resulting oligonucleotides are analyzed by MS in a manner that provides the relative proportion of triplet repeat oligonucleotides in seconds per sample. This assay enables swift and reproducible detection of expanded CGG alleles using a single blood spot and in principle is suitable for large scale studies and newborn screening. Moreover, this analytical scheme establishes a unique new intersection of MS with molecular biology, with potential for significant interdisciplinary impact.