APOE Genotyping for Alzheimer's Risk

APOE genotype, the strongest genetic risk factor for Alzheimer’s disease with effect on memory over the adult life course [42 (link)], was assessed through a venous blood sample. Genetic determination of APOE allelic status was performed using a polymerase chain reaction (PCR)-based assay designed with the MassARRAY Assay Design 4.0 software (Agena Bioscience, San Diego, CA, USA). The initial PCR step involved 45 cycles with an annealing temperature of 56 °C. The PCR products were treated with shrimp alkaline phosphatase for 15 min at 37 °C and denatured at 85 °C for 5 min. The final iPLEX extension step involved 40 cycles of lots of five cycles between 52 °C and 85 °C. The resulting iPLEX extension products were desalted using SpectroCLEAN resin (SEQUENOM, San Diego, CA, USA), then spotted on SpectroCHIPs GenII (SEQUENOM, San Diego, CA, USA) and analysed with the MassARRAY Analyser Compact MALDI-TOF MS (Agena Bioscience, San Diego, CA, USA). Participants were categorised as possessing at least one copy of the ε4 allele from the APOE (ε4 carrier) or no copies of this polymorphism (non-ε4).