Simple no-chromatography procedure for amine-reactive Eu³⁺ luminescent chelates optimal for bioconjugation

Lanthanide ions luminescence has long life time enabling highly sensitive detection in time-gated mode. The synthesis of reactive lanthanide probes for covalent labeling of the objects of interest is cumbersome task due to the large size of the probes, complex multi-step procedures and the presence of sensitive groups, which often prevents introduction of reactive cross-linking functions optimal for conjugation. We suggest simple synthetic protocol for luminescent europium chelates based on serendipitous reaction yielding acylating compounds, whose reactivity is comparable to that of commonly used N-hydroxysuccinimide (NHS) esters. The probes react with proteins at pH 7.0 within several minutes at ambient temperature displaying high coupling efficiency. The resulting conjugates survive electrophoretic separation under denaturing conditions, which makes the labels useful in proteomic studies that rely on high detection sensitivity.