Yes, they can – testing Alex’s cells for pluripotency

In the last blog we gave an overview on how to characterize iPS cells. Today, we dive in and explain how we test the cells for pluripotency, the potential to turn into different cell types of the human body.

 

Alex: Hi, it's been a while, but it's summer and I heard Lauren has accepted a new position!

Birgitt: Yes, we'll miss Lauren and we wish her the best of luck for her future.

Alex: So what's next?

Birgitt: I'll explain how we test your cells to see if they have the potential to show "yes we can" "Change"  into other cells. To test proteins and markers in the cell you need to visualize them. This can be done with a method called immunostaining. Immunostaining is an antibody-based method that detects specific proteins on the surface or inside of the cells. The picture below is an example of how we can immunostain the cells. We use an antibody specific for a certain marker on the cell, called epitope. Once this antibody binds to the epitope we then incubate with a fluorescently labeled antibody that binds to the first antibody. We can then visualize this fluorescence with a fluorescent microscope and take the colorful images you will see in the next few blogs. 

More info about immunostaining here

Alex: Hmm sounds a lot like 2008. I posted some pretty fluorescence pictures on my Facebook page that you shared with me in the past. I am looking forward to understanding more about these immunostaining techniques.

Birgitt: We used three different antibodies to detect whether or not your stem cells are pluripotent. The three markers we used were OCT4, SOX2, and SSEA4, these are proteins that are expressed in pluripotent cells and therefore called pluripotency markers. These are abbreviations for protein names octamer-binding transcription factor 4 (OCT4), sex determining region Y-box 2 (SOX2), and stage-specific embryonic 4 (SSEA4). Since these protein names are so complicated we usually only use the abbreviations.

See the results in the three images below.  The blue color is the nuclear stain or counterstain which shows where the nucleus of the cell is located. The green fluorescence corresponds one of the pluripotency markers.

iPS colony stained with SSEA-4 antibody

iPS colony stained with OCT4 antibody

iPS colony stained with SOX2 antibody

Alex: I hope we can talk about this more next time and I am also very excited to come out to San Francisco for the Festival of Genomics on November 5^th, 2015 to chat with you on stage about my Parkinson’s disease, stem cells,  and more (http://www.festivalofgenomicscalifornia.com/).

Birgitt: I am thrilled to have the opportunity to talk with you on stage, can't wait! And maybe the topic of pluripotency will be reelected in the next post.