{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,4,2]],"date-time":"2026-04-02T17:35:12Z","timestamp":1775151312368,"version":"3.50.1"},"reference-count":22,"publisher":"Wiley","issue":"3","license":[{"start":{"date-parts":[[2001,6,1]],"date-time":"2001-06-01T00:00:00Z","timestamp":991353600000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"funder":[{"DOI":"10.13039\/100004440","name":"Wellcome Trust","doi-asserted-by":"publisher","id":[{"id":"10.13039\/100004440","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Immunol Cell Biol"],"published-print":{"date-parts":[[2001,6]]},"abstract":"<jats:p>Real\u2010time quantitative reverse transcriptase\u2013polymerase chain reaction (RT\u2013PCR) is the method of choice for rapid and reproducible measurements of cytokine or growth factor expression in small samples. Fluorescence detection methods for monitoring real\u2010time PCR include fluorogenic probes labelled with reporter and quencher dyes, such as Taqman probes or Molecular Beacons and the dsDNA\u2010binding dye SYBR Green I. Fluorogenic (Taqman) probes for a range of human and rat cytokines and growth factors were tested for sensitivity and compared with an assay for SYBR Green I quantification using real\u2010time fluorescence monitoring (PE Applied Biosystems Model 7700 sequence detector). SYBR Green I detection involved analysis of the melting temperature of the PCR product and measurement of fluorescence at the optimum temperature. Fluorogenic probes provided sensitive and reproducible detection of targets that ranged from low (&lt;10 copies\/reaction) to high (&gt;10<jats:sup>7<\/jats:sup> copies\/ reaction) expression. SYBR Green I gave reproducible quantification when the target gene was expressed at moderate to high levels (\u22651000 copies\/reaction), but did not give consistently reproducible quantification when the target gene was expressed at low levels. Although optimization of melting temperature improved the specificity of SYBR Green I detection, in our hands it did not equal the reproducible sensitivity and specificity of fluorogenic probes. The latter method is the first choice for measurement of low\u2010level gene expression, although SYBR Green I is a simple and reproducible means to quantify genes that are expressed at moderate to high levels.<\/jats:p>","DOI":"10.1046\/j.1440-1711.2001.01002.x","type":"journal-article","created":{"date-parts":[[2003,3,12]],"date-time":"2003-03-12T00:43:03Z","timestamp":1047429783000},"page":"213-221","source":"Crossref","is-referenced-by-count":239,"title":["Real\u2010time reverse transcriptase\u2013polymerase chain reaction (RT\u2013PCR) for measurement of cytokine and growth factor mRNA expression with fluorogenic probes or SYBR Green I"],"prefix":"10.1111","volume":"79","author":[{"given":"Jian Lin","family":"Yin","sequence":"first","affiliation":[{"name":"Department of Renal Medicine, Royal Prince Alfred Hospital  New South Wales Australia"}]},{"given":"Nicholas A","family":"Shackel","sequence":"additional","affiliation":[{"name":"Centenary Institute, Royal Prince Alfred Hospital and University of Sydney  New South Wales Australia"}]},{"given":"Amany","family":"Zekry","sequence":"additional","affiliation":[{"name":"Centenary Institute, Royal Prince Alfred Hospital and University of Sydney  New South Wales Australia"}]},{"given":"Peter H","family":"McGuinness","sequence":"additional","affiliation":[{"name":"Centenary Institute, Royal Prince Alfred Hospital and University of Sydney  New South Wales Australia"}]},{"given":"Craig","family":"Richards","sequence":"additional","affiliation":[{"name":"Centenary Institute, Royal Prince Alfred Hospital and University of Sydney  New South Wales Australia"}]},{"given":"Karien","family":"Van Der Putten","sequence":"additional","affiliation":[{"name":"Centenary Institute, Royal Prince Alfred Hospital and University of Sydney  New South Wales Australia"}]},{"given":"Geoffrey W","family":"McCaughan","sequence":"additional","affiliation":[{"name":"Centenary Institute, Royal Prince Alfred Hospital and University of Sydney  New South Wales Australia"}]},{"given":"Josette M","family":"Eris","sequence":"additional","affiliation":[{"name":"Department of Renal Medicine, Royal Prince Alfred Hospital  New South Wales Australia"}]},{"given":"G Alex","family":"Bishop","sequence":"additional","affiliation":[{"name":"Centenary Institute, Royal Prince Alfred Hospital and University of Sydney  New South Wales Australia"}]}],"member":"311","published-online":{"date-parts":[[2001,6]]},"reference":[{"key":"e_1_2_7_2_1","doi-asserted-by":"publisher","DOI":"10.1111\/j.1600-065X.1991.tb00583.x"},{"key":"e_1_2_7_3_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.87.7.2725"},{"key":"e_1_2_7_4_1","doi-asserted-by":"publisher","DOI":"10.1097\/00007890-199206000-00023"},{"key":"e_1_2_7_5_1","doi-asserted-by":"publisher","DOI":"10.1084\/jem.174.2.493"},{"key":"e_1_2_7_6_1","doi-asserted-by":"publisher","DOI":"10.1038\/icb.1997.19"},{"key":"e_1_2_7_7_1","doi-asserted-by":"publisher","DOI":"10.1101\/gr.6.10.995"},{"key":"e_1_2_7_8_1","first-page":"954","article-title":"Quantification of low\u2010copy transcripts by continuous SYBR Green I monitoring during amplification","volume":"24","author":"Morrison TB","year":"1998","journal-title":"Biotechniques"},{"key":"e_1_2_7_9_1","doi-asserted-by":"publisher","DOI":"10.1038\/nbt0396-303"},{"key":"e_1_2_7_10_1","doi-asserted-by":"publisher","DOI":"10.1046\/j.1365-3083.1999.00549.x"},{"key":"e_1_2_7_11_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0022-1759(97)00188-9"},{"key":"e_1_2_7_12_1","doi-asserted-by":"publisher","DOI":"10.1016\/0003-2697(87)90021-2"},{"key":"e_1_2_7_13_1","doi-asserted-by":"publisher","DOI":"10.1016\/0966-3274(93)90033-5"},{"key":"e_1_2_7_14_1","doi-asserted-by":"publisher","DOI":"10.1097\/00007890-199405150-00011"},{"key":"e_1_2_7_15_1","doi-asserted-by":"publisher","DOI":"10.1038\/sj.bjc.6690552"},{"key":"e_1_2_7_16_1","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.88.16.7276"},{"key":"e_1_2_7_17_1","doi-asserted-by":"publisher","DOI":"10.1126\/science.7683443"},{"key":"e_1_2_7_18_1","doi-asserted-by":"publisher","DOI":"10.1006\/mcpr.1999.0284"},{"key":"e_1_2_7_19_1","doi-asserted-by":"publisher","DOI":"10.1101\/gr.4.4.219"},{"key":"e_1_2_7_20_1","doi-asserted-by":"publisher","DOI":"10.1006\/cyto.1998.0426"},{"key":"e_1_2_7_21_1","doi-asserted-by":"publisher","DOI":"10.1007\/s002239900717"},{"key":"e_1_2_7_22_1","doi-asserted-by":"publisher","DOI":"10.1016\/S0165-2427(99)00100-2"},{"key":"e_1_2_7_23_1","doi-asserted-by":"publisher","DOI":"10.1101\/gr.1.1.70"}],"container-title":["Immunology &amp; 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