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The workflow of single-cell expression profiling using quantitative real-time PCR

Journal article
Authors Anders Ståhlberg
Mikael Kubista
Published in Expert Review of Molecular Diagnostics
Volume 14
Issue 3
Pages 323-331
ISSN 1473-7159
Publication year 2014
Published at Sahlgrenska Cancer Center
Institute of Biomedicine, Department of Pathology
Pages 323-331
Language en
Links dx.doi.org/10.1586/14737159.2014.90...
Keywords single-cell workflow, gene expression profiling, RT-qPCR, single-cell analysis, preamplification, single-, GUIDELINES MINIMUM INFORMATION, MESSENGER-RNA LEVELS, GENE, DNA, QUANTIFICATION, PUBLICATION, MOLECULES, DISEASE, CANCER, FLOW
Subject categories Pathology

Abstract

Biological material is heterogeneous and when exposed to stimuli the various cells present respond differently. Much of the complexity can be eliminated by disintegrating the sample, studying the cells one by one. Single-cell profiling reveals responses that go unnoticed when classical samples are studied. New cell types and cell subtypes may be found and relevant pathways and expression networks can be identified. The most powerful technique for single-cell expression profiling is currently quantitative reverse transcription real-time PCR (RT-qPCR). A robust RT-qPCR workflow for highly sensitive and specific measurements in high-throughput and a reasonable degree of multiplexing has been developed for targeting mRNAs, but also microRNAs, non-coding RNAs and most recently also proteins. We review the current state of the art of single-cell expression profiling and present also the improvements and developments expected in the next 5 years.

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