Volume 12 Issue 8, August 2017

Hydrogels, networks of water-swollen polymers, are being exploited for the local delivery of cells and biologically relevant molecules. Loebel et al. describe the preparation of supramolecular hydrogels and their characterization.

This protocol describes how to construct luciferase probes that are targeted to the mitochondrial matrix or the outer surface of the plasma membrane. These probes can be used to measure ATP concentrations in different cellular compartments.

Bhatia et al. describe how to prepare giant unilamellar vesicles by using complex lipid mixtures and proteins (Na⁺/K⁺-ATPase) at physiological conditions; the protocol includes subsequent imaging of lateral nanoscale structures of the membrane.

Sun et al. describe how to image and quantify mitophagy in both living cells and tissues, using the pH-sensitive fluorescent reporter mt-Keima. This protocol provides information for analysis by both confocal and super-resolution microscopy.

Fluorescent peptides with excellent target specificity are useful imaging probes. This protocol describes the synthesis of a tryptophan-based fluorogenic amino acid (Fmoc-Trp(C₂-BODIPY)-OH) and its incorporation into peptides for live-cell imaging.

This protocol describes how to make and use a gelatin-based hypoxia-inducible hydrogel to inject or embed tissue or cells. This enables the cellular responses to controllable hypoxic gradients to be assessed in vitro and in vivo, e.g., in mice.

Chen, Cang and colleagues describe how to characterize intracellular ion channels using a manual patch-clamp technique on enlarged organelles such as endolysosomes.

This protocol describes DAP-seq, a transcription-factor binding site discovery assay that can be used to produce cistrome and epicistrome maps for any organism.

This protocol provides a computational pipeline for accurate detection and grouping of viral sequences from microbiome samples. The approach uses a set of viral protein families as bait for identifying viral sequences directly from metagenomic assemblies.

This protocol describes the deep-scale analysis of the blood plasma proteome. By combining abundant protein depletion, sample multiplexing with isobaric labeling and fractionation, this enables rapid quantification of >4,500 plasma proteins.

Glycoengineering of IgG antibodies and glycosite-specific antibody–drug conjugates is an important tool for enhancing their therapeutic efficacy. Tang et al. describe a protocol for efficient and homogeneous chemoenzymatic antibody glycoengineering.