Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
5
pubmed:dateCreated
2003-5-19
pubmed:abstractText
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 cytokine expression with accuracy and no post-PCR manipulation.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
http://linkedlifedata.com/resource/pubmed/chemical/Actins, http://linkedlifedata.com/resource/pubmed/chemical/Cytokines, http://linkedlifedata.com/resource/pubmed/chemical/Fluorescent Dyes, http://linkedlifedata.com/resource/pubmed/chemical/Indicators and Reagents, http://linkedlifedata.com/resource/pubmed/chemical/Interferon-gamma, http://linkedlifedata.com/resource/pubmed/chemical/Interleukin-12, http://linkedlifedata.com/resource/pubmed/chemical/Interleukin-12 Subunit p40, http://linkedlifedata.com/resource/pubmed/chemical/Interleukin-2, http://linkedlifedata.com/resource/pubmed/chemical/Ionomycin, http://linkedlifedata.com/resource/pubmed/chemical/Organic Chemicals, http://linkedlifedata.com/resource/pubmed/chemical/Protein Subunits, http://linkedlifedata.com/resource/pubmed/chemical/RNA, Messenger, http://linkedlifedata.com/resource/pubmed/chemical/SYBR Green I, http://linkedlifedata.com/resource/pubmed/chemical/Tetradecanoylphorbol Acetate, http://linkedlifedata.com/resource/pubmed/chemical/Transforming Growth Factor beta
pubmed:status
MEDLINE
pubmed:month
May
pubmed:issn
0300-9475
pubmed:author
pubmed:issnType
Print
pubmed:volume
57
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
439-45
pubmed:dateRevised
2008-11-21
pubmed:meshHeading
pubmed-meshheading:12753500-Actins, pubmed-meshheading:12753500-Animals, pubmed-meshheading:12753500-Computer Systems, pubmed-meshheading:12753500-Cost-Benefit Analysis, pubmed-meshheading:12753500-Costs and Cost Analysis, pubmed-meshheading:12753500-Cytokines, pubmed-meshheading:12753500-Female, pubmed-meshheading:12753500-Fluorescent Dyes, pubmed-meshheading:12753500-Gene Expression Profiling, pubmed-meshheading:12753500-Indicators and Reagents, pubmed-meshheading:12753500-Interferon-gamma, pubmed-meshheading:12753500-Interleukin-12, pubmed-meshheading:12753500-Interleukin-12 Subunit p40, pubmed-meshheading:12753500-Interleukin-2, pubmed-meshheading:12753500-Ionomycin, pubmed-meshheading:12753500-Lymphocyte Activation, pubmed-meshheading:12753500-Mice, pubmed-meshheading:12753500-Mice, Inbred BALB C, pubmed-meshheading:12753500-Oligonucleotide Array Sequence Analysis, pubmed-meshheading:12753500-Organic Chemicals, pubmed-meshheading:12753500-Polymerase Chain Reaction, pubmed-meshheading:12753500-Protein Subunits, pubmed-meshheading:12753500-RNA, Messenger, pubmed-meshheading:12753500-Sensitivity and Specificity, pubmed-meshheading:12753500-Spleen, pubmed-meshheading:12753500-T-Lymphocytes, pubmed-meshheading:12753500-Tetradecanoylphorbol Acetate, pubmed-meshheading:12753500-Transforming Growth Factor beta
pubmed:year
2003
pubmed:articleTitle
Quantification of cytokine gene expression using an economical real-time polymerase chain reaction method based on SYBR Green I.
pubmed:affiliation
Departamento de Inmunología, Escuela Nacional de Ciencias Biológicas, Instituto Politécnico Nacional, México D F, México.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't, Evaluation Studies