Parameters involved in the conversion of real-time PCR assays from the ABI prism 7700 to the Cepheid SmartCycler II.

Clin Biochem

Department of Pathology, Dartmouth Medical School, Dartmouth-Hitchcock Medical Center, One Medical Center Drive, Lebanon, NH 03756, USA.

Published: February 2005

Objective: We examined several critical parameters that must be optimized when converting between the ABI Prism 7700 real-time PCR platform and the Cepheid SmartCycler II while using the same primer and probe sequences.

Design And Methods: A lyophilized master mix, MgCl(2) concentration, PCR cycling conditions, and ramp times were evaluated.

Results: Optimization of each parameter, including use of the OmniMix HS-lyophilized beads, 6 mM MgCl(2) concentration, changes in PCR cycling parameters, and increased ramp time were necessary to convert this real time PCR assay to a new platform.

Conclusion: We conclude that careful consideration of several analytical parameters can result in a smooth transition of assays between real time PCR platforms.

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Source
http://dx.doi.org/10.1016/j.clinbiochem.2004.10.009DOI Listing

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