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      Genomics Protocols 

      Transcriptional Profiling of Small Samples in the Central Nervous System

      Humana Press

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          Most cited references 27

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          Generalized singular value decomposition for comparative analysis of genome-scale expression data sets of two different organisms.

          We describe a comparative mathematical framework for two genome-scale expression data sets. This framework formulates expression as superposition of the effects of regulatory programs, biological processes, and experimental artifacts common to both data sets, as well as those that are exclusive to one data set or the other, by using generalized singular value decomposition. This framework enables comparative reconstruction and classification of the genes and arrays of both data sets. We illustrate this framework with a comparison of yeast and human cell-cycle expression data sets.
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            Making and reading microarrays.

            There are a variety of options for making microarrays and obtaining microarray data. Here, we describe the building and use of two microarray facilities in academic settings. In addition to specifying technical detail, we comment on the advantages and disadvantages of components and approaches, and provide a protocol for hybridization. The fact that we are now making and using microarrays to answer biological questions demonstrates that the technology can be implemented in a university environment.
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              In situ detection of apoptotic nuclei in the substantia nigra compacta of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-treated mice using terminal deoxynucleotidyl transferase labelling and acridine orange staining.

               N.A Tatton,  S.J. Kish (1997)
              The neurotoxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) was used to generate a dose-dependent cell death of dopaminergic nigral neurons in the C57B1 mouse. Mice were injected with a total cumulative dose of 150 mg/kg of MPTP delivered over five days and killed at different time points both during and after the toxin injections. Two independent histological methods were used to determine whether the dopaminergic nigral neurons died via an apoptotic mechanism. In situ end-labelling with terminal deoxynucleotidyl transferase was used on paraformaldehyde-fixed, serial, frozen sections to identity cells with double-stranded DNA breaks. Apoptotic cell death was found to be initiated within 72 h of the first injection of the neurotoxin and peaked 24 h after the final MPTP injection. The metachromatic fluorochrome, Acridine Orange, was used on alternate sections to provide structural confirmation of the nuclear chromatin "clumping" considered to be representative of apoptosis. Confocal laser imaging combined with deconvolution techniques was used to resolve the fluorescent signal emitted by the in situ Acridine Orange-DNA complexes. The number of Acridine Orange-stained nuclei demonstrating chromatin clumping was identical to that of the positive in situ end-labelled nuclei observed over a 25 day period. Based upon these two independent methods of assessing apoptosis in situ, we conclude that a 150 mg/kg dose of MPTP can elicit apoptotic cell death in nigral dopaminergic neurons of the C57B1 mouse.
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                Author and book information

                Book
                978-1-58829-871-3
                978-1-59745-188-8
                2008
                10.1007/978-1-59745-188-8
                Book Chapter
                2008
                : 147-158
                10.1007/978-1-59745-188-8_10

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