General


“Animal behavior weaves itself throughout the tapestry of biology: It is sparked when neurons fire in response to stimuli in the external world, it forms the interactions that lead to reproduction and genetic propagation, and it enhances complex group function.”Michael J. Ryan and Walter Wilczynski

Animal BehaviorThis clip from An Introduction to Animal Behavior, our newest book, reminds us that the all animal behaviors—mate choice, social bonding, migration, cooperation, conflict, and aggression—have a biological basis involving internal processes (e.g., genetics, neurobiology, and physiology) and external factors (e.g., environment and social surroundings).  In the book, Drs. Michael J. Ryan and Walter Wilczynski connect animal behaviors to both evolutionary considerations and mechanistic processes, providing an integrated view of why animals behave the way they do.  Drs. Ryan and Wilczynski describe classic behavioral studies in the context of new research findings in areas as diverse as endocrinology, phylogenetics, development, anatomy and physiology, molecular genetics, and neurobiology.  Extensively illustrated, An Introduction to Animal Behavior provides a well-thought-out introduction to the complexity of animal behavior, and should appeal to advanced undergraduates, graduate students, and professional scientists.

For more information about the book, click here.

This will be my last post on Bench Marks as I am leaving CSHL Press for a new opportunity at Oxford University Press. I’ve had a good 10 year run at Cold Spring Harbor and leave with warm memories and a much greater understanding of the world of publishing.

Bench Marks will live on though, and there are plans to do all sorts of interesting things with the blog, so stay tuned. Maria Smit will now be taking over as the managing editor of Cold Spring Harbor Protocols and will be continuing to keep this blog updated with news of new techniques and interesting articles.

Thanks to all who have read this blog and the readers and authors who have made Cold Spring Harbor Protocols into such a huge success. I’ll still be around at my other blogging gig at The Scholarly Kitchen so look for me there.

The ability to image living cells and tissues has revolutionized our understanding of many biological processes. Being able to visualize what’s happening in these time lapse movies at an ultrastructural level would make them even more informative. But commonly used fluorescent proteins like GFP are not directly visible by electron microscopy (EM). Fluorescent nanoparticles or quantum dots can be visualized by EM, but targeting these to cytoplasmic proteins in living cells remains difficult. One method which does allow correlation between light microscopy and EM is fluorescence photoconversion, where observation of the fluorescent label causes the deposition of a reaction product that can be rendered electron-dense and directly visualized by EM. In the January issue of Cold Spring Harbor Protocols, Mark Ellisman and colleagues from UCSD’s National Center for Microscopy and Imaging Research provide a set of articles for this method in combination with a class of genetically encoded peptide tags that can be labeled in living cells by fluorophores bearing two appropriately spaced arsenic atoms (biarsenicals).

Correlated Live Cell Light and Electron Microscopy Using Tetracysteine Tags and Biarsenicals provides an overview of the technique, and protocols are provided for Labeling Tetracysteine-Tagged Proteins with Biarsenical Dyes for Live Cell Imaging and Fluorescence Photoconversion of Biarsenical-Labeled Cells for Correlated Electron Microscopy (EM).

New technologies and methods are spurring a renaissance in the study of organogenesis. Organogenesis, essentially the process through which a group of cells becomes a functioning organ, has important connections to biological processes at the cellular and developmental levels, and its study offers great potential for medical treatments through tissue engineering approaches. The January issue of Cold Spring Harbor Protocols features a method from Washington University’s Hila Barak and Scott Boyle for Organ Culture and Immunostaining of Mouse Embryonic Kidneys. The kidney is particularly interesting as it also serves as a model for branching morphogenesis. The protocol describes the isolation, culture and fluorescent immunostaining of mouse embryonic kidneys. As one of January’s featured articles, the protocol is freely available to subscribers and nonsubscribers alike.

The “Brainbow” strategy was originally developed in mice, as a system for labeling neurons in a variety of different colors, allowing one to follow multiple cells regardless of their proximity. Brainbow uses a construct that carries sequences for red, blue and green fluorescent proteins in tandem array, with two pairs of lox sites flanking the first two fluorescent proteins. Recombination occurs in the presence of the Cre recombinase, and one gets a variety of outcomes, resulting in the production of a red, blue or green label. When more than one copy of the Brainbow cassette exists within a cell, the primary tones can be mixed, providing more possible color combinations. This provides a powerful platform for studying neuronal morphology and cell movements. In the January issue of Cold Spring Harbor Protocols, Alex Schier and colleagues offer Multicolor Brainbow Imaging in Zebrafish. This protocol translates the system for use in zebrafish, which offer the advantages of easy visualization of transparent embryos and efficient generation of labeled subjects.

Cap analysis gene expression (CAGE) is a method used to discover new promoters and for quantifying gene activity, providing data essential for studies of regulatory gene networks. But CAGE requires large amounts of RNA, which are often not obtainable from rare specimens. In the January issue of Cold Spring Harbor Protocols Piero Carninci and colleagues from the RIKEN Yokohama Institute’s Omics Science Center present NanoCAGE: A High-Resolution Technique to Discover and Interrogate Cell Transcriptomes, a method that can capture information from as little as 10 nanograms of total RNA. The protocol describes how to rapidly prepare nanoCAGE libraries which can be sequenced with high sensitivity. As one of January’s featured articles, the protocol is freely available to subscribers and non-subscribers alike.

How do you see something smaller than the wavelength of light itself? Fluorescence microscopy is the most common optical technique used for visualizing cellular functions. The latest techniques allow labeling of specific organelles and proteins with molecular precision. But conventional microscopy cannot resolve objects closer than 200 nanometers at the focal plane. Many subcellular structures and groups of proteins occur on the 10 nanometer scale. A true understanding of cellular physiology requires new superresolution methods.

Of these methods, PALM (Photoactivated Localization Microscopy) provides the highest shown resolution in biological samples (approximately 10 nanometers) and allows for the assessment of individual molecules. In the December issue of Cold Spring Harbor Protocols, Oregon Health Science University’s Haining Zhong presents Photoactivated Localization Microscopy (PALM): An Optical Technique for Achieving ~10-nm Resolution. The article provides an overview of the basic principles of PALM, its implementation and the potential applications in neuroscience.

Array tomography is a volumetric microscopy method based on physical serial sectioning. Ultrathin sections of tissue are cut using an ultramicrotome, and attached in order to a glass coverslip. These coverslips are then stained as desired and imaged. The resulting two-dimensional image tiles can then be reconstructed computationally into three-dimensional volume images for visualization and quantitative analysis. The thin sections allow for rapid staining and imaging and the array format allows much of the process to be automated. In the November issue of Cold Spring Harbor Protocols, Stephen J. Smith and colleagues from Stanford University present Array Tomography: High-Resolution Three-Dimensional Immunofluorescence, a guide to this technique that allows for visualizing previously inaccessible features of tissue structure and molecular architecture. As one of November’s featured articles, it is freely accessible to subscribers and non-subscribers alike.

N-terminalomics is a high-throughput strategy for identifying proteins by selectively enriching for and sequencing their N-terminal peptides by mass spectrometry. In the November issue of Cold Spring Harbor Protocols, Samie Jaffrey and colleagues from Cornell University present a newly-developed N-terminalomic approach, N-CLAP (N-terminalomics by Chemical Labeling of the alpha-Amine of Proteins). N-CLAP: Global Profiling of N-Termini by Chemoselective Labeling of the alpha-Amine of Proteins describes the use of Edman chemistry to modify all of the amines in proteins, followed by the generation of a new unmodified amine at the N-terminus after the removal of the first amino acid by peptide bond cleavage. The alpha-amine at the protein N-terminus is labeled with a cleavable biotin affinity tag, which facilitates the downstream purification of the N-terminal peptides. Peptides are eluted by cleaving the biotin affinity tag and identified by tandem mass spectrometry (MS/MS). N-CLAP can be used for the identification of signaling peptides for mature proteins as well as for global profiling of cleavage events that occur during cell signaling, such as apoptosis.

Imaging has rapidly become a defining tool of the current era in biological research. But finding the right method and optimizing it for data collection can be a daunting process, even for an established imaging laboratory. Cold Spring Harbor Protocols is one of the world’s leading sources for detailed technical instruction for implementation of imaging methods, and the November issue features articles detailing standard and cutting-edge laboratory techniques.

The confocal microscope is a workhorse of the modern life science laboratory. Its popularity stems from its ability to permit volume objects to be imaged and rendered in three dimensions. But the confocal microscope itself does not produce three-dimensional images; in fact, it only images very thin sections of a specimen that lie within its focal region. To produce a three-dimensional image, a series of thin optical sections are collected, and computer processing is used to combine them into a volumetric rendering. In the first of November’s featured articles, Spinning-Disk Microscopy Systems, Oxford University’s Tony Wilson reviews the many methods for producing optical sections, of which the confocal optical system is just one. He also describes a number of convenient methods of implementation that can lead to, among other things, real-time image formation. The paper, like all our featured articles, is freely available to subscribers and non-subscribers alike.

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