Methods in molecular biology
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The heat/capsaicin sensitization and intradermal capsaicin injection models are safe and noninvasive paradigms to generate stable, long-lasting, and reproducible injury capable of producing an area of both primary and secondary hyperalgesia. Risk of skin injury is substantially reduced since lower levels of thermal and chemical irritation produce long-lasting cutaneous hyperalgesia. ⋯ Unlike the heat/capsaicin sensitization model, intradermal capsaicin results in a brief painful stimulus followed by a long lasting area of secondary hyperalgesia. The intradermal injection of capsaicin results in a transient, intense stinging sensation at the site of injection (e.g. heat allodynia) followed by a persistent area of secondary tactile allodynia.
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Limited understanding of the cell biology of the breast and breast cancer hampers our ability to develop new therapeutic approaches. Mouse models of mammary gland development and tumourigenesis are key to developing new insights into the biology of both the normal and diseased tissues. Recent advances have enabled the isolation, molecular characterisation and functional analysis of mouse mammary epithelial cell subpopulations from the normal gland, including subpopulations enriched for stem cell behaviour. Application of these techniques both to the normal mammary gland and to tumour models will promote a better understanding of the nature of the different epithelial cell types in the mammary gland, the origins of mammary tumours and the role of tumour stem cells.
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This book is part of the Methods in Molecular Biology series, and provides a general overview of computational approaches used in proteome research. In this chapter, we give an overview of the scope of the book in terms of current proteomics experimental techniques and the reasons why computational approaches are needed. We then give a summary of each chapter, which together provide a picture of the state of the art in proteome bioinformatics research.
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Amplifying and sequencing DNA after bisulfite treatment of genomic DNA reveals the methylation state of cytosine residues at the highest resolution possible. However, a thorough analysis is required for statistical evaluation of methylation at all sites in each genomic region. Several software tools were developed to assist in quantitative evaluation of bisulfite sequencing data from complex methylation patterns occurring in plants. ⋯ CyMATE is also able to perform a quality control of sequences and to detect redundancy among individual clones. The software is able to reveal methylation patterns on complementary strands by handling data from hairpin bisulfite sequencing. The tool is freely available for non-commercial use at http://www.cymate.org .
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The retinol carrier retinol-binding protein (RBP) forms in blood a complex with the thyroid hormone carrier transthyretin (TTR). The interactions of retinoid-RBP complexes, as well as of unliganded RBP, with TTR can be investigated by means of fluorescence anisotropy. RBP represents the prototypic lipocalin, in the internal cavity of which the retinol molecule is accommodated. ⋯ The fluorescence anisotropy technique is also suitable to study the interaction of TTR with apoRBP and RBP in complex with non-fluorescent retinoids. In the latter cases, the fluorescence signal is provided by a fluorescent probe covalently linked to TTR rather than by RBP-bound retinol. We report here on the preparation of recombinant human RBP and TTR, the covalent labeling of TTR with the fluorescent dansyl probe, and fluorescence anisotropy titrations for RBP and TTR.