We have screened exon 12 of the low density lipoprotein (LDL) receptor gene from 46 familial hypercholesterolemia (FH) heterozygotes and 64 normolipidemic controls for two polymorphisms, HincII, which is caused by a T to C substitution at base 1773, and a C to T transition at base 1725, by using single strand conformation polymorphism (SSCP) analysis. Our results indicate that polymorphism at base 1725, previously reported as very rare from a Japanese sample, is quite frequent in the Spanish population and that it is closely linked to the presence of the HincII site (HincII+). Thus, both polymorphisms constitute a system of three alleles, typed HincII- C1725, HincII+ C1725, and HincII+ T1725, whose frequencies in the FH sample were 0.489, 0.347, and 0.164, respectively. No significant differences were found in the allele frequencies between the FH and control samples. This three-allelic polymorphic system provides a higher information content (PIC value) than the HincII RFLP alone (0.537 versus 0.373, respectively); therefore, it is an extremely useful marker for linkage analysis of FH in Caucasian populations.