Tennielle Ludwig

WiCell Research Institute

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Focus on stem cell quality: Improving rigor and reproducibility through characterization

As stem cell scientists, the quality of our research is directly related to the quality of the hPSC materials used. Poor quality cells can impact reproducibility, jeopardize results, waste time, and drain resources. In screening materials submitted to the WiCell Stem Cell Bank, we have identified a substantial and concerning variability in cell quality. Further in-depth analysis of a decade of karyotypic data has allowed us to identify specific areas of recurrent karyotypic instability previously unknown. These results highlight the need for improved testing strategies and standards.

As of this abstract, more than 1600 cell lines have been deposited with the WiCell Stem Cell Bank for banking and characterization by 31 providing laboratories. The vast majority of these cell lines were generated through grant-funded projects as a resource for the larger scientific community, and reportedly screened prior to submission. Various testing strategies were used, and available characterization information was provided to WiCell for reference. To date, nearly 800 of these lines have been independently tested by WiCell for thaw viability, genetic stability (karyotype), identity via short tandem repeat (STR) analysis, sterility (bacteria and fungus), and mycoplasma. Of the hPSC lines examined, more than one-third of WiCell screened cell lines failed this routine quality testing. While there were failures across all tests, the majority of cell lines failed due to unexpected abnormal karyotype.

Stem cell lines deposited with the WiCell Stem Cell Bank are karyotyped internally, and WiCell Characterization additionally performs genetic testing for outside organizations. We performed a retrospective analysis on karyotype data collected over the course of a decade, (including more than 15,000 hPSC cultures). This analysis enabled us to identify striking shifts in relative frequencies of recurrent abnormalities; namely, dramatically increasing rates of chromosomes 1 and 20 gains at the expense of chromosome 12 gains. Additionally, we identified the minimal amplicon for all chromosome 1q gains as chromosomal band (segment) 1q32.1, suggesting that this region harbors the driver gene(s) that give this recurrent aberration its advantage in culture.

Overall, these results show that quality screening strategies in use today are variable, and largely insufficient. Based on this data, we can assume that a substantial percentage of materials used in investigator laboratories have unidentified quality issues that will impact research. The reliability and reproducibility of data gained through experimentation is dependent on maintaining normal, consistent cell cultures. Changes in genetic composition can have dramatic impacts on cell function, and therefore experimental results. This underscores the need for routine testing prior to initiating and following studies, particularly genetic analysis to assure cell line stability.


Tenneille Ludwig currently serves as the Director of the WiCell Stem Cell Bank overseeing the banking, distribution, and core services operations at WiCell. Dr. Ludwig obtained her Bachelor's and Master’s degrees from Washington State University prior to completing a Ph.D. in embryology and developmental biology with a minor in bioethics from UW–Madison in 2001. Her subsequent work in the laboratory of Dr. James Thomson (2001-2007) focused primarily on the optimization of cell culture conditions and resulted in the development of the first defined, feeder-independent culture system for human embryonic stem cells (TeSR/mTeSR). Dr. Ludwig has served as a Scientific Advisor to multiple boards, including the International Stem Cell Forum Ethics Working Party, and collegiate training programs. Currently, she is a member of the Stem Cell and Regenerative Medicine Center and the ESCRO Committee at the University of Wisconsin-Madison, and serves on the Steering Committee for both the International Stem Cell Banking Forum (ISCBF) and the International Stem Cell Initiative Genetics and Epigenetics Study Group. Her primary focus remains collaborating within these groups to refine consensus standards for banking, characterization, and distribution of research and cGMP grade materials, and working with investigators to support the advancement of stem cell technologies through clinical trial.

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