Alex's stem cells pass the practice test

In the last blog we talked about how we bank your cells. Today, we will give you an overview of how to make sure that the cell we have are actually pluripotent stem cells, a process we call characterization.

Lauren: We use four methods to characterize your cells. In the image below you can see a summary of these methods. Two methods test the pluripotency of the newly derived cells and the other two methods make sure that the DNA was not damaged or changed during the reprogramming process.

Summary of characterization process

Alex: I know we have talked about pluripotency before, but can you remind me what that is?

Lauren: Pluripotency refers to the ability of the cells to give rise to all of the cell types that make up the human body. The first way we test pluripotency is by staining the cells for certain markers. All cell types have markers on the surface of their cells and within the cells. These markers are specific to each type of cell so that we can distinguish different cells from one another. We use multiple markers that are specific for pluripotency to be sure that the cells are indeed pluripotent.

Alex: Well that seems fairly straightforward. How else do you test for pluripotency?

Lauren: Another way that we test for pluripotency is to spontaneously differentiate these stem cells. The spontaneous differentiation begins by forming embryoid bodies. Embryoid bodies are 3-D cell aggregates as shown in the picture below. The cells are no longer adherent to the bottom of the dish but are floating in the media. Embryoid bodies are meant to mimic the early development of an embryo. This stage primes the stem cells and gets them ready to differentiate into all cell types. We use specific cell markers to confirm that the stem cells can differentiate into various tissue types.

Example of embryoid bodies

Alex: I did not realize that so much work went into these processes. What else needs to be done in order to call my newly made stem cells "induced pluripotent stem cells"?

Lauren: The next two methods of characterization ensure that the DNA within the cells is not damaged and that the virus has not "sneaked into" the DNA by the reprogramming process. The first method is called a karyotype which shows the number and shape of chromosomes (packaged DNA) of the cell. For humans, a normal karyotype consists of 22 pairs of chromosomes and the two sex chromosomes X and/or Y. The image below is an example of how a normal karyotype looks like.

Example of a normal karyotype, 46 XY

Alex: I hope my karyotype comes out normal. What is the next step?

Lauren: To make sure that the genetic material of these reprogrammed cells has not been modified, we measure the virus in the cells which we used to deliver the reprogramming factors.  After 10 to 12 passages, we should not detect the virus anymore. We can detect the virus by an amplification method that will be explain in more detail later.

Alex: Thank you for the overview. I am looking forward to learning more about each of these methods and seeing if my cells pass these tests. It is a bit nerve racking almost like passing a practice driving test.

Lauren: Indeed, I have quite nervous about some of the test results of your cells. In the next few blogs we will go into more detail about each of these methods and show the results in you cells. The first process we will discuss is staining for the pluripotency cell markers.