We have developed a rapid, nonradioactive screening test enabling the simultaneous analysis of three lowdensity Iipoprotein receptor (LDLR) gene mutations (DIS4N, D206E, and V408M), which together account for familial hypercholesterolemla (FH) in -90% of the South African Afrikaner population. The assay is designed so that FH patients, negative for these founder-related mutations (found in descendants of European settlers), subsequently can be screened for unknown mutations in the mutation-rich exon 4 of the LDLR gene. Our screening assay consists of two steps: (1) multiplex allele-specific PCR amplification of exons 4 and 9, and (2) simultaneous analysis of single-and double-strand conformational polymorphisms in exon 4 by vertical electrophoresis on low crosslinked polyacrylamide gels. The simplicity, specificity, and versatility of the multiplex assay makes it an ideal system for routine screening of FH mutations in large population samples.

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