A Validation Study of the Applied Biosystems[TM] GlobalFiler[TM] PCR Amplification Kit on the Applied Biosystems[TM] 3500xL Genetic Analyzer

The main focus of this internship was the validation of the Applied Biosystems[TM] GlobalFiler[TM] PCR Amplification Kit on the Applied Biosystems[TM] 3500xL Genetic Analyzer. Internal validations are essential in accredited forensic laboratories, as it is required by Standard 8 of the FBI Quality Assurance Standards (2011) that new procedures, instruments and kits, have to be validated prior to their implementation into casework. According to the SWGDAM guidelines, the validation study has to include several studies; known and non-probative evidence samples or mock evidence samples, sensitivity and stochastic studies, precision and accuracy studies, a concordance study, mixture studies, a degradation study and a contamination assessment. At the Harris County Institute of Forensic Sciences (HCIFS), the validation of the Applied BiosystemsTM 3500xL Genetic Analyzer was conducted in the Genetics Laboratory. The 3500xL Genetic Analyzer has numerous enhanced capabilities over the older instrument that HCIFS currently uses for casework; the Applied BiosystemsTM 3130xL Genetic Analyzer. The 3500xL can analyze six dyes, has RFID, has 24 capillaries, and a solid-state long-life laser. The samples were extracted and purified for use on the 3500xL via QIAgen or the Organic method. The Quant Trio Kit, and the GlobalFilerTM PCR Amplification Kit were also used in the process. After injecting the samples on the 3500xL, they were analyzed appropriately for each study. 28 cycles for amplification and a 24 second injection time were used. An analytical threshold of 50 RFU for blue, 65 RFU for yellow, 45 RFU for green, 55 RFU for red and 60 RFU for purple were used, with a stochastic threshold of 400 RFUs. An optimal DNA amount of 0.5ng was also used. The results of each study showed that the 3500xL to give reliable and reproducible results, and the instrument performed as expected.