TCO-PEG4-acid is a non-activated PEG derivative containing a TCO moiety and a terminal carboxylic acid. The terminal carboxylic acid can be reacted with primary amine groups in the presence of activators (e.g. EDC, or DCC) to form a stable amide bond. This reagent can react with tetrazine-containing compounds to form a stable covalent bond without the catalysis of Cu or elevated temperatures. The inverse-electron demand Diels-Alder cycloaddition reaction of TCO with tetrazines is the fastest bioorthogonal reaction with exceptional selectivity. The hydrophilic PEG spacer increases solubility in aqueous media.