NCRAD - The National Cell Repository for Alzheimer's Disease
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Processing of Samples

DNA and Cell LinesPlasma and SerumRNA

How DNA and Cell Line Samples Are Stored at NCRAD

  1. The main function of the National Cell Repository for Alzheimer’s Disease involves collecting, storing, and distributing genetic material for Alzheimer’s disease (AD) research. Our goal is to provide the scientific community with a valuable resource for obtaining the genetic material needed to study families with AD. Researchers from around the world may request cell lines or DNA, the material that is used in genetic research. All DNA and cell lines are always de-identified (no identifying information such as name or date of birth is made available) before being distributed. Research on these samples does not provide any diagnostic testing information.

  2. The process of banking (storing) DNA and cell lines begins when a blood sample is received from a member of a family having AD or serious memory loss.

  3. The sample is taken to the lab and assigned a unique kit number. The kit number and information about the participant is recorded in a secured database.

  4. The sample is then assigned a unique bar code. The bar code number is entered into a logbook along with the unique kit number. These numbers are checked by several technicians for accuracy.

  5. The blood sample is then placed in a machine and spun to separate the sample into three main layers: the red blood cell layer, the plasma layer, and the buffy coat, which contains the white blood cells. The white blood cells are needed to establish cell lines and obtain DNA.


    6a. To establish cell lines, the white blood cells are placed into two separate flasks along with a solution that allows permanent cell growth. The cells are incubated at 37ºC (body temperature) anywhere from three weeks to three months for continuous cell division.   6b. To isolate DNA, the white blood cells are washed and spun at a high speed, enabling the cells to cluster together.
             
    7a. The cell-containing solutions are then transferred into two larger flasks for further cell growth. It takes approximately one week for the cells to divide to the desired number. The cells are checked throughout this process to ensure that they are growing properly.   7b. The clusters of white blood cells are placed in a solution containing an enzyme that degrades unnecessary cell components. The solution is stored at 98ºF overnight. Tee cells are split open during this time while the DNA stays intact.
             
    8a. The cells are then placed in a plastic cryo-vial along with a cryo-preservative. Each vial holds approximately 1 milliliter of solution containing 1x107 (10000000 cells).   8b. Next, the DNA is separated from the cell components. This is accomplished by adding a salt solution and spinning the DNA and cell components at a high speed.
             
    9a. The cells are gradually cooled to freezing temperatures and then placed in a tank of liquid nitrogen. The slow freeze prevents damage to the cell line and takes place in a controlled-rate freezer.   9b. The DNA is isolated and transferred to another tube, which now contains the desired DNA without the unnecessary cell components. A form of alcohol is added to the solution and interacts with the DNA. This allows the DNA to gather together and become visible.
             
    10a. The frozen cells are stored in a tank filled with liquid nitrogen at -316ºF. Cells can be preserved this way indefinitely and thawed at any time for additional cell growing. This culture and storage process is necessary for immortalizing our participants’ cells for ongoing genetic research in AD.   10b. The DNA is then transferred to a small tube containing a solution that preserves the genetic material for future studies. The DNA is stored for future use in a freezer set at -94ººF. If a sample does not grow, the individual who provided the sample may be contacted and asked to provide another blood sample.
  1. Since 1991, NCRAD has collected more than 12000 samples from individuals and families. These samples and the clinical and family history information from these individuals have been used by researchers to understand the genetics of AD. Over the past 18 years, over 29000 samples have been distributed to researchers study AD. More than 80 different investigators from around the world have requested samples from NCRAD. These researchers have published 177 papers about AD and dementia. These papers have helped to better understand the genetics of early onset and late onset AD as well as other types of dementia.

    The cell lines and DNA are extremely valuable for AD research. We greatly appreciate all the support from the families who participate in NCRAD.

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