Selenoprotein Structure and Function, Volume 662 in the Methods in Enzymology series, highlights new advances in the field, with this new volume presenting interesting chapters written by an international board of authors. Chapters in this new release include Identification of Selenoprotein O substrates using a biotinylated ATP analog, Selenium-encoded isotopic signature targeted profiling, Designing tRNASec variants for efficient selenocysteine incorporation using Sec-specific reporters, Preparation of selenoprotein S by chemical ligation, Examining xCT-mediated selenium uptake and selenoprotein production capacity in cells, SecMS analysis of selenoprotein with selenocysteine insertion sequence and beyond, Selenocysteine substitutions in thiyl radical enzymes, and much more. Additional chapters cover Recombinant selenoprotein expression in E. coli based upon the redefinition of a UAG codon in an RF1-depleted host strain, Metabolic labeling with radioactive selenium in zebrafish, Low pH isoTOP-ABPP to identify selenocysteines, Expression of selenoproteins via genetic code expansion in mammalian cells, Alpha-methyl selenocysteine as a tool for the study of selenoproteins, Selective selenol fluorescent probes: design, synthesis, structural determinants, and biological applications, and much more.