Abstract
► Luminescent avidin probes carrying multiple Eu and Tb chelates were obtained. ► Detection sensitivity of the probes was in 10⁻¹⁴–10⁻¹⁵ M range. ► The probes were used for labeling and time-resolved imaging of living cells.
Lanthanide ion luminescence has a long lifetime enabling highly sensitive detection in time-gated mode. The sensitivity can be further increased by using multiple luminescent labels attached to a carrier molecule, which can be conjugated to an object of interest. We found that up to 30 lanthanide chelates can be attached to avidin creating highly bright constructs. These constructs with Eu³⁺ chelates display synergistic effect that enhance the brightness of heavily modified samples, while the opposite effect was observed for Tb³⁺ chelates thereby significantly reducing their light emission. This undesirable quenching of Tb³⁺ luminophores was completely suppressed by the introduction of an aromatic spacer between the chelate and the protein attachment site. The estimated detection limit for the conjugates is in the 10⁻¹⁴–10⁻¹⁵ M range. We demonstrated a high sensitivity for the new probes by using them to label living cells of bacterial and mammalian origin.