A total of 74 non-pregnant women with a previous episode of thrombosis were investigated for activated protein C (APC) resistance in the aPTT-based and factor IXa-X-based assays and for the presence of mutations in all APC-cleavage sites in the heavy chains of factor V and factor VIII. DNA fragments were amplified with the polymerase chain reaction (PCR) and those encoding for the Arg-306 and Arg-506 (factor V) and for Arg-740 (factor VIII) cleavage sites were subjected to restriction enzyme analysis. DNA fragments of 29 selected patients corresponding to the Arg-306 and Arg-679 cleavage sites in factor V, and to the Arg-336 and Arg-562 cleavage sites in factor VIII were sequenced. APC resistance was found in 40 cases, using the aPTT-based assay and in 35, using the factor IXa-X-based assay (23 patients were APC resistant in both assays), whereas 22 individuals had a normal response to APC. Forty-three patients carried Arg-506 to Gln mutation in factor V. No other polymorphism or mutation was found in the genes for factors V or VIII in the vicinity of the APC cleavage sites. It was concluded that the difference in response to APC in the two assays may not be explained by the presence of mutations in the APC cleavage sites of factor V and factor VIII in this group of patients. The data do not exclude the presence of mutations elsewhere in the factor V or factor VIII genes.