Maxwell dna ffpe kit

This study included 157 previously untreated PMBCL patients from collaborating centers in 5 countries. Additional 83 samples with available RNA and known ZNF217 mutation status were studied [20 (link), 22 (link)] (Supplementary Table 1).
In general, we included patients above the age of 18, if their clinical presentation (dominated by a mediastinal mass) and a reviewed histology report including estimation of tumor content (G.O., I.A., or T.V.) were consistent with the diagnosis of PMBCL according to the WHO [23 (link)]. Tumor purity was calculated using the ALL-FIT algorithm [24 (link)]. For FFPE tumor tissue, DNA was extracted using the Maxwell DNA-FFPE Kit (Promega, Madison, WI, USA). For fresh frozen tumor tissue (FF), genomic DNA was extracted using the Maxwell 16 Tissue DNA Purification Kit. RNA was isolated using the Maxwell® RSC RNA FFPE Kit.
The study was conducted in accordance with the Declaration of Helsinki. The protocol was approved by the ethics review committee of the Charité - Universitätsmedizin Berlin, and of every participating center. All patients gave written informed consent.

All human studies were performed under the Institutional Review Board approval at the University of Wisconsin. Pickhardt et al2 (link) serially monitored 306 polyps ranging in starting size from 6 to 9 mm by CT colonography. Per cent volumetric grow rate per year was determined as previously described.2 (link) Of the 306 with known growth fates, 48 resected polyps were selected based on amount of remaining formalin-fixed paraffin-embedded (FFPE) tissue and from a variety of growth fates. These selected tumours were removed from 36 asymptomatic patients identified at normal colorectal cancer screening from the University of Wisconsin Hospital and Clinics as well as the Walter Reed National Military Medical Center. These individuals had a mean age of 57±8 years and 28 (78%) were male. DNA was isolated from FFPE tissue scraped from 5 µm sections using the Maxwell DNA FFPE Kit (Promega, Madison, Wisconsin, USA) and eluted into a volume of 30 µL of buffer following the manufacturer's instructions.

Low-frequency variants for which commercial primers were available were validated with TaqMan Mutation Detection Assays (Thermo Fisher) (see online supplementary figure S1). Mutation detection was performed according to assay instructions. Briefly, FFPE tissue was microdissected from multiple regions of each tumour (see online supplementary figure S1A–C) under a dissection microscope and DNA was purified using the Maxwell DNA FFPE Kit (Promega). Samples were run in duplicate or triplicate as per manufacturer's instructions to determine the presence of or the frequency of the variant DNA, respectively. The qPCR reactions were run on Bio-Rad CFX96 Real-Time PCR and data were analysed using the Mutation Detector software (Thermo Fisher, last revised April 2012). Variants that fell below our variant calling cut-offs for sequencing, but that were validated by qPCR were included in the dataset. This included the KRAS variant for polyp PF24 and the removal of the CTNNB1 variant in PF11.