Methods in molecular biology
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In a normal spontaneous menstrual cycle, the luteal phase is characterized by the production and secretion of estradiol (E) and progesterone (P) from the corpus luteum (CL) in an episodic manner. The steroidogenesis of the CL is dependent on continued tonic luteinizing hormone (LH) secretion (Fritz and Speroff, Clinical gynecologic endocrinology and infertility, 8th edn. Wolters Kluwer, Lippincott Williams & Wilkins, Philadelphia, 2011). ⋯ Progesterone concentrations normally rise sharply after ovulation, reaching a peak approximately 8 days after the LH surge. Since the secretion of E and P during the luteal phase is episodic and correlates closely with LH pulses, relatively low mid-luteal progesterone levels can be found in the course of a totally normal luteal phase (Fritz and Speroff, Clinical gynecologic endocrinology and infertility, 8th edn. Wolters Kluwer, Lippincott Williams & Wilkins, Philadelphia, 2011).
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Chondroitin sulphate proteoglycans (CSPGs) are one of the major families of inhibitory extracellular matrix molecules in the central nervous system. The expression of various CSPGs is strong during early nervous system development; however, it is downregulated during maturation and up-regulated again after nervous system injury. In vivo injection of an enzyme called chondroitinase ABC, which removes the inhibitory chondroitin sulphate chains on the CSPGs, in the injured area promotes both the regeneration and plasticity of the neurons. Here, we describe the method of in vivo injection of the chondroitinase ABC into the cortex of adult rat brain and the histochemical method to assess the successfulness of the digestion.
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As the field of proteomics shifts from qualitative identification of protein "subfractions" to quantitative comparison of proteins from complex biological samples, it is apparent that the number of approaches for quantitation can be daunting for the result-oriented biologist. There have been many recent reviews on quantitative proteomic approaches, discussing the strengths and limitations of each. ⋯ Here we present a detailed bioinformatics workflow for one of the simplest, most pervasive quantitative approach-spectral counting. The informatics and statistical workflow detailed here includes newly available freeware, such as SePro and PatternLab which post-process data according to false discovery rate parameters, and statistically model the data to detect differences and trends.
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Experimental spinal cord injury (SCI) can maintain the continuity of the spinal cord, as in the contusion (e.g., weight-fall) or compression models, or not, when there is a partial or a complete transection. The majority of acute human SCI is not followed by complete transection, but there is a combination of contusion, compression, and possibly partial transection. ⋯ This lesion was established by our group and represents a simple, reliable, and inexpensive clip compression model with functional and morphological reproducibility. In this chapter we describe, step by step, the protocol of this experimental SCI.
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In this first, introductory chapter, it is intended to summarize from a methodological point of view the state of the art in plant proteomics, focusing on mass spectrometry-based strategies. Thus, this chapter is mainly directed at beginners or at those trying to get into the field, rather than at those with real experience or a long trajectory in plant proteomics research. The different alternative workflows, methods, techniques, and protocols from the experimental design to the data analysis will be briefly commented, with cross references to previous monographs and reviews, as well as to the rest of the book chapters. ⋯ Furthermore, and even more important, we should move to data validation through other -omics or classical biochemical strategies, in an attempt to get a deeper, real, and more accurate view and understanding of cell biology. In the modern Systems Biology concept, proteomics must be considered as a part of a global, multidisciplinary approach. Making biological sense of a proteomics experiment requires a proper experimental design, data validation, interpretation, and publication policy.