跳至主要内容

A Combination of DNA-peptide Probes and Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS): A Quasi-Targeted Proteomics Approach for Multiplexed MicroRNA Quantification

文献作者：Feifei Xu, Weixian Zhou, Jianxiang Cao, Qingqing Xu, Dechen Jiang and Yun Chen   《THERANOSTICS》

ABSTRACTS
The distorted and unique expression of microRNAs (miRNAs) in cancer makes them an attractive source of biomarker. There is much evidence indicating that a panel of miRNAs, termed “miRNA fingerprints”, is more specific and informative than an individual miRNA as biomarker. Thus, multiplex assays for simultaneous quantification of multiple miRNAs could be more potent in clinical practice. However, current available assays normally require pre-enrichment, amplification and labeling steps, and most of them are semi-quantitative or lack of multiplexing capability. In this study, we developed a quasi-targeted proteomics assay for multiplexed miRNA quantification by a combination of DNA-peptide probes and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Specifically, the signal of target miRNAs (i.e., miR-21, miR-let7a, miR-200c, miR-125a and miR-15b) was converted into the mass response of reporter peptides by hybridization of miRNAs with DNA-peptide probes and subsequent tryptic digestion to release the peptides. After a careful optimization of conditions related to binding, conjugation, hybridization and multiple reaction monitoring (MRM) detection, the assay was validated for each miRNA and the limit of quantification (LOQ) for all the miRNAs can achieve 1 pM. Moreover, crosstalk between DNA-peptide probes in multiplex assay was sophisticatedly evaluated. Using this quasi-targeted proteomics assay, the level of target miRNAs was determined in 3 human breast cell lines and 36 matched pairs of breast tissue samples. Finally, simplex assay and qRT-PCR were also performed for a comparison. This approach grafts the strategy of targeted proteomics into miRNA quantification and may offer a new way for multiplexed miRNA profiling.
KEY WORDS
Quasi-Targeted Proteomics, MicroRNAs, Multiplex Assay, Reporter Peptides, DNA-peptide Probes, Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS).
SCREENSHOT

RELATED PRODUCTS
Peptides including maleimide-modified substrate peptide, reporter peptide and internal standard containing stable-isotope labeled amino acids were developed by ChinaPeptides Co., Ltd. (Shanghai, China).
CHAINING
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5562220/
标签：

评论

发表评论

此博客中的热门博文

Image-guided stem cells with functionalized self-assembling peptide nanofibers for treatment of acute myocardial infarction in a mouse model

文献作者：Xiao Li, Ying-Ying Chen, Xiu-Mei Wang, Kai Gao, Yun-Zhou Gao, Jian Cao, Zhuo-Li Zhang, Jing Lei, Zheng-Yu Jin, and Yi-Ning Wang     《American Journal of Translational Research》 ABSTRACTS Aim: To investigate the survival of bone marrow mesenchymal stem cells (BMSCs) and the therapeutic effect for acute myocardial infarction (AMI) after co-transplantation with the functionalized self-assembling peptide nanofiber RAD/PRG or RAD/KLT. Methods: AMI of balb/c mice was induced. Mice were randomly divided into four groups, and received treatment of phosphate buffered saline (PBS) (Group A), GFP/Fluc-BMSCs (Group B), GFP/Fluc-BMSCs + RAD/PRG (Group C), and GFP/Fluc-BMSCs + RAD/KLT (Group D), respectively. Bioluminescence imaging (BLI) was performed on day 1 (d-1), d-4, d-7, d-10, and d-13 after transplantation. Magnetic resonance imaging (MRI) was performed at baseline (d-4 before transplantation) and d-29 after treatment. Mice were euthanized on d-29 following tr...
发表评论
阅读全文

The therapeutic effect of anti-CD52 treatment in murine experimental autoimmune encephalomyelitis is associated with altered IL-33 and ST2 expression levels

文献作者：Mark Barbour, Rachel Wood, Shehla U.Hridi, Chelsey Wilson, Grant McKay, Trevor J.Bushell, Hui-Rong Jiang   《Journal of Neuroimmunology》 ABSTRACTS Experimental autoimmune encephalomyelitis  (EAE) mice were administered with murine anti-CD52  antibody  to investigate its therapeutic effect and whether the treatment modulates IL-33 and ST2 expression. EAE severity and  central nervous system  (CNS) inflammation were reduced following the treatment, which was accompanied by peripheral  T and B lymphocyte  depletion and reduced production of various  cytokines  including IL-33, while sST2 was increased. In spinal cords of EAE mice, while the number of IL-33 +  cells remained unchanged, the extracellular level of IL-33 protein was significantly reduced in anti-CD52 antibody treated mice compared with controls. Furthermore the number of ST2 +  cells in the spinal cord of treated EAE mice was  downregulated ...
发表评论
阅读全文

Near-infrared light-activated red-emitting upconverting nanoplatform for T1-weighted magnetic resonance imaging and photodynamic therapy

Author：Xiang-long Tang, Jun Wu, Ben-lan Lin, Sheng Cui, Hong-mei Liu, Ru-tong Yu, Xiao-dong Shen, Ting-wei Wang, Wei Xia     《Acta Biomaterialia》 ABSTRACTS Photodynamic therapy (PDT) has increasingly become an efficient and attractive cancer treatment modality based on reactive oxygen species (ROS) that can induce tumor death after  irradiation   with ultraviolet or visible light. Herein, to overcome the limited tissue penetration in traditional PDT, a novel near-infrared (NIR) light-activated NaScF4: 40% Yb, 2% Er@CaF2  upconversion  nanoparticle   (rUCNP) is successfully designed and synthesized.  Chlorin   e6, a  photosensitizer   and a chelating agent for Mn2+, is loaded into  human serum albumin   (HSA) that further conjugates onto rUCNPs. To increase the ability to target glioma tumor, an acyclic  Arg–Gly–Asp peptide   (cRGDyK) is linked to rUCNPs@HSA(Ce6-Mn). This nanoplatform enables e...
发表评论
阅读全文