APR 17, 2014 11:00 AM PDT

A Novel Method of Active Paraffin Removal

Sponsored by: Covaris, Covaris
Speaker
  • Principal Scientist, Covaris, Inc.
    Biography
      Dr. Hamid Khoja brings years of experience in assay, protocol, and kit development in the field of molecular biology. Recently, Dr. Khoja has helped advance FFPE DNA and RNA extraction by utilizing Covaris' patented Adaptive Focused Acoustics Technology to develop a robust protocol and the truXTRAC™ purification kits. Previously, as a research scientist, Dr. Khoja developed several kits and high throughput assays for internal research initiatives, including branched DNA assays for oncogene quantification, Mass spectrometer based assays for CFTR and other disease mutation analysis, kits for ChIP-ChIP assays and the truChIP™ Chromatin Shearing reagent kits optimizing chromatin shearing for ChIP applications.
       
       

    Abstract

     

    Formalin Fixation and Paraffin Embedding (FFPE) of tissue samples is a routine practice by pathologists; the technique enables high quality preservation of structural information. However, this method was developed to facilitate histological analysis rather than for the new molecular analysis approaches which are emerging in clinical oncology settings (such as whole genome sequencing). For example, the chemical crosslinks and the paraffin matrix of FFPE samples are a very significant challenge for the robust, high yield extraction of nucleic acids. To enable reliable access to the molecular information of clinical samples, we developed a novel, robust method for FFPE samples using Covaris Adaptive Focused Acoustics (AFA). Efficient, high yield, and high quality extraction of DNA from FFPE tissues for molecular analysis applications is possible by the novel use of Covaris' patented Adaptive Focused Acoustics™ (AFA) technology. The focused, short wavelength, acoustic field generated by Covaris AFA allows for active paraffin removal without the use of organic solvents. It also enables complete tissue rehydration, the effective digestion of proteins, and the efficient release of DNA. The simplified workflow from sample to purified DNA is less than 3 hours (less than 10 minutes hands-on time). We will present whole genome sequencing results from truXTRAC™ FFPE extracted DNA, and RNA-Seq results from truXTRAC FFPE extracted RNA. In a comparative study, truXTRAC showed both greater yield and greater coverage depth across the genome than other commercial extraction methods. More importantly, coverage uniformity with the truXTRAC kit was very similar to that of DNA extracted from matched fresh frozen tissues in gene rich regions of the genome. The technique also allows for the parallel processing of up to 8 FFPE samples in a SBS format 96 well plate. We present data showing highly reproducible results while processing a large number of FFPE samples.


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