Nature Protocols 4, 1709 (2009). doi:10.1038/nprot.2009.159

Authors: Mardas Daneshian, Sonja von Aulock & Thomas Hartung

We present an internationally validated protocol for the evaluation of pyrogenic contaminations using human whole blood. In the in vitro pyrogen test (IPT) the sample is incubated with fresh or cryopreserved human whole blood, and the proinflammatory cytokine interleukin-1β (IL-1β) is detected by enzyme-linked

Nature Protocols 4, 1722 (2009). doi:10.1038/nprot.2009.183

Authors: Susan M Gribble, Bee Ling Ng, Elena Prigmore, Tomas Fitzgerald & Nigel P Carter

Array painting is a technique that uses microarray technology to rapidly map chromosome translocation breakpoints. Previous methods to map translocation breakpoints have used fluorescence in situ hybridization (FISH) and have consequently been labor-intensive, time-consuming and restricted to the low breakpoint resolution imposed by the

Nature Protocols 4, 1737 (2009). doi:10.1038/nprot.2009.185

Authors: Anne Limbourg, Thomas Korff, L Christian Napp, Wolfgang Schaper, Helmut Drexler & Florian P Limbourg

Blood vessel growth in adult organisms involves the following two fundamental processes: angiogenesis, the proliferation and extension of capillary networks; and arteriogenesis, the growth of functional arteries. We provide a protocol for the evaluation of postnatal arteriogenesis and angiogenesis in a mouse model of hind-limb

Nature Protocols 4, 1749 (2009). doi:10.1038/nprot.2009.190

Author: Maria A Serrat

Fluorescent microspheres are commonly used to assess bone blood supply in large animals, but the technique is not widely used in smaller mammals, as traditional methods such as reference blood sampling, ventilation and catheterization are not easily applied. This protocol describes a viable alternative for

Nature Protocols 4, 1759 (2009). doi:10.1038/nprot.2009.174

Authors: Nokyoung Park, Jason S Kahn, Edward J Rice, Mark R Hartman, Hisakage Funabashi, Jianfeng Xu, Soong Ho Um & Dan Luo

Cell-free systems represent a promising approach to quickly and easily produce preparative amounts of proteins. However, it is still challenging to obtain high volumetric yields (>mg ml−1) of proteins from the present cell-free systems. This protocol presents a cell-free protein synthesis method using

Nature Protocols 4, 1771 (2009). doi:10.1038/nprot.2009.184

Authors: Daniel J Turner & Matthew E Hurles

When combined with haplotype fusion PCR (HF-PCR), ligation haplotyping is a robust, high-throughput method for empirical determination of haplotypes, which can be applied to assaying both sequence and structural variation over long distances. Unlike alternative approaches to haplotype determination, such as allele-specific PCR and long

Nature Protocols 4, 1784 (2009). doi:10.1038/nprot.2009.188

Authors: Chang C Liu, Susan E Cellitti, Bernhard H Geierstanger & Peter G Schultz

Tyrosine sulfation is an important post-translational modification that occurs in higher eukaryotes and is involved in cell–cell communication, viral entry and adhesion. We describe a protocol for the heterologous expression of selectively tyrosine-sulfated proteins in Escherichia coli through the use of an expanded genetic

Nature Protocols 4, 1790 (2009). doi:10.1038/nprot.2009.189

Authors: Andrea Cossarizza, Roberta Ferraresi, Leonarda Troiano, Erika Roat, Lara Gibellini, Linda Bertoncelli, Milena Nasi & Marcello Pinti

Reactive oxygen species (ROS) are continuously produced in the cell as a consequence of aerobic metabolism, and are controlled by several antioxidant mechanisms. An accurate measurement of ROS is essential to evaluate the redox status of the cell, or the effects of molecules with the

Nature Protocols 4, 1798 (2009). doi:10.1038/nprot.2009.191

Authors: Florence Debacq-Chainiaux, Jorge D Erusalimsky, Judith Campisi & Olivier Toussaint

Normal cells can permanently lose the ability to proliferate when challenged by potentially oncogenic stress, a process termed cellular senescence. Senescence-associated beta-galactosidase (SA-βgal) activity, detectable at pH 6.0, permits the identification of senescent cells in culture and mammalian tissues. Here we describe first a cytochemical

Nature Protocols 4, 1807 (2009). doi:10.1038/nprot.2009.192

Authors: Ceri A Morris, Elizabeth Benson & Helen White-Cooper

We describe a whole-mount RNA in situ hybridization (ISH) method optimized for detection of the cellular and subcellular distributions of specific mRNA within Drosophila testes and male genital tract. Digoxygenin (dig)-labeled antisense RNA probes are in vitro transcribed from a template synthesized

Nature Protocols 4, 1820 (2009). doi:10.1038/nprot.2009.194

Authors: Janine Mok, Hogune Im & Michael Snyder

Herein, we describe a protocol for the global identification of in vitro substrates targeted by protein kinases using protein microarray technology. Large numbers of fusion proteins tagged at their carboxy-termini are purified in 96-well format and spotted in duplicate onto amino-silane-coated slides in a

Nature Protocols 4, 1828 (2009). doi:10.1038/nprot.2009.198

Authors: Azar Radfar, Darío Méndez, Carlos Moneriz, María Linares, Patricia Marín-García, Antonio Puyet, Amalia Diez & José M Bautista

This protocol describes a method for preparing cultures of Plasmodium falciparum synchronized at any intraerythrocytic stage. Using this method, around 60% parasitized cells may be obtained. On the basis of Trager and Jensen's original continuous culture method, our approach relies on the use of