eBook: Cell and Gene Therapy - 23

Use of Quantitative Mycoplasma
DNA Controls in Evaluating the
Sensitivity of Molecular-based
Detection Systems
Novel controls offer an effective and efficient means for the
detection of mycoplasma contamination
Stephen King, MS; Brian Chase, PhD; Afshin Sohrabi, PhD; Maria Mayda, PhD;
Kurt Langenbach, PhD; Cara N. Wilder, PhD
Abstract
Novel molecular-based methods offer an effective
and efficient means for the detection of mycoplasma
contamination in cell cultures. However,
for these methods to become widely accepted in
research laboratories, they must demonstrate sensitivity,
particularly with regard to the limit of detection.
Here, we describe the use of quantitative
mycoplasma DNA controls in the evaluation of two
molecular-based detection systems.
Introduction
Mycoplasma contamination affects roughly 1535%
of continuous cell cultures, resulting in a number
of deleterious effects including the induction
of chromosomal abnormalities, the disruption of
DNA and RNA synthesis, and the inhibition of both
cell metabolism and growth rate.1-3
Historically, the
recommended mycoplasma testing protocol for research
laboratories included the use of traditional
culture-based techniques and DNA staining with
fluorochromes.4
Despite the successful use of these
detection methods for many years, their limit of
detection remains undefined. Moreover, the entire
testing procedure is laborious, costly, and difficult
to interpret. However, before an alternative method,
such as a molecular-based detection system,
can become widely accepted and implemented,
this method must exhibit sensitivity with regard to
the limit of detection.
To this end, ATCC has designed quantitative mycoplasma
DNA controls for use in testing and calibration
in ISO 17025 accredited laboratories, inclusivity/exclusivity
testing, establishing limits of
23
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