Artus ebv qs rgq kit

Plasma EBV DNA concentrations were evaluated before treatment (pre-EBV), at the 5^th week of the radiation course (mid-EBV) and 3 months after the completion of radiation treatment (post-EBV). We elected to test the mid-EBV in the 5^th week of IMRT because it was the best time to perform re-simulation and in accordance with Leung’s study which tested mid-EBV at completion of 4 weeks of radiation therapy.21 (link) The EBV nucleic acids were purified from the plasma samples using the QIAsymphony SP in combination with the QIAsymphony DSP Virus/Pathogen Midi Kit (QIAGEN, Germany) using the manufacturer’s recommended protocol. After extraction, the eluates in the 96-microwell plates were transferred to the module for assembly with the master mix (QIAGEN artus EBV QS RGQ kit) by the instrument. The aliquoted reactions were subsequently put in a Rotor-Gene Q. The amplification parameters were as follows: 95°C for 10 min and 45 cycles of 95°C for 15 s, 65°C for 30 s, and 72°C for 20 s. The plasma DNA samples were quantified for EBV DNA using an RTQ-PCR system targeting the BamHI-W fragment region of the EBV genome. A plasma EBV DNA concentration of < 316 copies/ml was defined as an undetectable level in our institution. Note that in the following section, values of 0 represent an undetectable plasma EBV DNA concentration.

Nucleic acid was extracted according to the manufacturer's instructions. Using the QIAamp DSP DNA Mini Kit (Qiagen) with QIAcube, 100 µL eluate was obtained from 200 µL WB. Next, 30 µL of either the artus CMV RG Kit (Qiagen) or artus EBV RG Kit (Qiagen) master mix was added to 20 µL of the 100 µL eluate, for a final reaction volume of 50 µL.
Using the QIAsymphony DNA Mini Kit with QIAsymphony SP, 90 µL eluate was obtained from 200 µL (300 µL minus 100 µL dead volume) WB. A total of 60 µL internal control (IC) plasmid plus either CMV or EBV buffer was added to each sample prior to extraction. For the QIAsymphony AS component, 30 µL of either the artus CMV QS-RGQ kit (Qiagen) or artus EBV QS-RGQ kit (Qiagen) master mix was added to 20 µL (of the 60 µL) eluate, for a final reaction volume of 50 µL (Table 1).