TY - JOUR ID - 103971 TI - Absolute quantification of murine interleukine-4, interleukine- 10 and interferon-γ gene transcripts using Real Time PCR JO - Archives of Razi Institute JA - ARI LA - en SN - 0365-3439 AU - Esmaelizad, M. AU - Jafari, S. AU - Rafiei, Alireza AD - Central Laboratory Department, Razi Vaccine and Serum Research Institute, Karaj, Iran AD - Central Laboratory Department, Razi Vaccine and Serum Research Institute, Karaj, Iran Molecular Cell Biology Research Center, Mazandaran University of Medical Sciences, Sari, Iran AD - Molecular Cell Biology Research Center, Mazandaran University of Medical Sciences, Sari, Iran Y1 - 2015 PY - 2015 VL - 70 IS - 2 SP - 119 EP - 125 KW - real time PCR KW - Cytokine KW - IL4 KW - IL10 KW - IFN KW - γ DO - 10.7508/ari.2015.02.007 N2 - The study of cytokines gene expression is quite important in various conditions of health and disease for the evaluation of clinical responses to new vaccination approaches. An absolute quantification is based on a calibration curve and production of standard controls to achieve more reliable results than in relative system. In this study we attempted to construct standard controls to evaluate the murine immune response. The number of 10 Balb/c mice immunized with hydatid cyst fluid subcutaneously with two week intervals to induce transcription of Th1 and Th2 related cytokines. Three Pairs of primers were designed to amplification of interleukine-4, interleukine-10 and interferon-γ by Oligo software. Partial sequences of three cytokine genes were cloned into pTZ57T vector. Three recombinant plasmids were purified and serial dilutions were prepared. Real time qPCR carried out using SYBR-Green I fluorescence dye and standard curves were provided by the 7500 ABI SDS software based on the exact concentration of dilutions and the amplification plots. Results showed that this method was able to evaluate the cytokines mRNA levels less than 0.01pg (~150 copy). We concluded that absolute real-time qPCR can be successfully applied to the quantification of antigen-induced cytokines. UR - https://archrazi.areeo.ac.ir/article_103971.html L1 - https://archrazi.areeo.ac.ir/article_103971_f4527141b0568e688a38bc55c49084ef.pdf ER -