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Pulse-chase methods offer powerful tools for following the evolution of a biological system over time, but are usually limited to ensemble measurements of the average behavior of very large numbers of cells. Here we describe three methods ranging from a true pulse-chase, through selective regional photoactivation, to pharmacological induction of an altered protein state, which can be applied to time-dependent studies at the single-cell level. These methods are exemplified by experimental protocols to follow region-selective nuclear envelope targeting of nascent phospholipids, a nascent nuclear lamin protein (lamin B1), and an immature lamin precursor (prelamin A).

Original publication

DOI

10.1007/978-1-4939-3530-7_10

Type

Journal article

Journal

Methods Mol Biol

Publication Date

2016

Volume

1411

Pages

159 - 176

Keywords

Nucleoplasmic reticulum, Photoactivatable fluorescent proteins, Pulse-chase, Secondary ion mass spectrometry, Single-cell assays, Stable isotope labeling, Super-resolution light microscopy