Optimization of small-scale sample preparation for high-throughput OpenArray analysis

Authors

  • Neeta A. Abraham Biogen
  • Anne C. Campbell Biogen
  • Warren D. Hirst Biogen
  • Catherine L. Nezich Biogen

DOI:

https://doi.org/10.14440/jbm.2021.339

Keywords:

Cells-to-CT, OpenArray, qPCR, RNeasy

Abstract

OpenArray is one of the most high-throughput qPCR platforms available but its efficiency can be limited by sample preparation methods that are slow and costly. To optimize the sample workflow for high-throughput qPCR processing by OpenArray, small-scale sample preparation methods were compared for compatibility with this system to build confidence in a method that maintains quality and accuracy while using less starting material and saving time and money. This study is the first to show that the Cells-to-CT kit can be used to prepare samples within the dynamic range of OpenArray directly from cultured cells in a single well of a 96-well plate when used together with a cDNA preamplification PCR step. Use of Cells-to-CT produced results of similar quality and accuracy to that of a preparation method using purified RNA in less than half the sample preparation time. While Cells-to-CT samples also exhibited slightly increased variance, which affects the ability of OpenArray to distinguish small differences in gene expression, overall gene expression mean results correlated well between small-scale methods. This work demonstrates that Cells-to-CT with preamplification can be used to reliably prepare samples for OpenArray analysis while saving time, money, and starting material.

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Published

2021-02-01

How to Cite

1.
Abraham NA, Campbell AC, Hirst WD, Nezich CL. Optimization of small-scale sample preparation for high-throughput OpenArray analysis. J Biol Methods [Internet]. 2021Feb.1 [cited 2021May8];8(1):e143. Available from: https://jbmethods.org/jbm/article/view/339

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