PMID: 12753500May 20, 2003Paper

Quantification of cytokine gene expression using an economical real-time polymerase chain reaction method based on SYBR Green I

Scandinavian Journal of Immunology
R Ramos-PayánI C E Estrada-Garcia

Abstract

Assessment of cytokine expression has become crucial to understand host responses to infections as well as autoimmunity. Several approaches including Northern blot, RNase protection assay and enzyme-linked immunosorbent assay have been used for this purpose, but they are time consuming, labour intense, and relatively large quantity of the samples is usually required. Recently, a technique termed real-time reverse transcriptase-polymerase chain reaction (RT-PCR) has been developed to determine genetic expression with great sensitivity and specificity; however, specialized instrumentation and costly reagents are usually needed. We aimed at using low-cost reagents for real-time PCR. This was achieved by adapting a conventional RT-PCR protocol to the quantitative real-time format, by the addition of the SYBR Green I reagent. We validated the approach by assessing the cytokine gene expression of murine splenocytes upon stimulation with phorbol 12-myristate 12-acetate (PMA)-ionomycin. The results using this technique were compared with those obtained with the well-established gene array method. We conclude that the use of the SYBR Green I reagent during real-time RT-PCR provides a highly specific and sensitive method to quantify cyto...Continue Reading

References

Apr 1, 1990·Proceedings of the National Academy of Sciences of the United States of America·G GillilandH F Bunn
Aug 1, 1996·BioTechniques·K Zimmermann, J W Mannhalter
Oct 1, 1996·Genome Research·C A HeidP M Williams
Oct 1, 1996·Genome Research·U E GibsonP M Williams
Jan 1, 1997·BioTechniques·C T WittwerR P Rasmussen
Mar 18, 1997·Proceedings of the National Academy of Sciences of the United States of America·R A HellerR W Davis
Feb 3, 1998·Nature Biotechnology·A Marshall, J Hodgson
Dec 23, 1998·Proceedings of the National Academy of Sciences of the United States of America·S D DerR H Silverman
Jan 23, 1999·BioTechniques·W M FreemanK E Vrana
Jun 3, 1999·Scandinavian Journal of Immunology·C HärtelH Klüter
Sep 7, 2001·The Journal of Immunology : Official Journal of the American Association of Immunologists·T ChtanovaC R Mackay
Oct 31, 2001·Journal of Immunological Methods·M L Garba, J A Frelinger

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Citations

May 22, 2008·Cancer Cell International·Marco A Velasco-VelázquezJuan J Mandoki
Feb 8, 2014·Developmental and Comparative Immunology·Patricia González-CanoPhilip J Griebel
Oct 17, 2007·FEMS Microbiology Letters·Blake Bextine, Beth Child
Aug 31, 2004·World Journal of Gastroenterology : WJG·Lin WangXue-Hai Tan
Apr 19, 2005·Expert Review of Molecular Diagnostics·Manit AryaHitendra R H Patel
Mar 1, 2005·Multiple Sclerosis : Clinical and Laboratory Research·Yu-Min HuangHans Link
Mar 7, 2007·The Journal of Immunology : Official Journal of the American Association of Immunologists·Aklile BerhanuWalter J Storkus
Feb 26, 2011·Clinical and Translational Science·Alan E RosenbachRonald G Tompkins
Aug 3, 2005·BioTechniques·Marisa L Wong, Juan F Medrano
Jul 8, 2021·Biologicals : Journal of the International Association of Biological Standardization·Vikas Kushwaha, Sukhbir Kaur
Sep 11, 2021·Biochimica Et Biophysica Acta. Biomembranes·Jun Hee OhYoonkyung Park

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