Time-resolved luminescence detection of peroxynitrite using a reactivity-based lanthanide probe

Abstract

Peroxynitrite (ONOO À ) is a powerful and short-lived oxidant formed in vivo, which can react with most biomolecules directly. To fully understand the roles of ONOO À in cell biology, improved methods for the selective detection and real-time analysis of ONOO À are needed. We present a water-soluble, luminescent europium(III) probe for the rapid and sensitive detection of peroxynitrite in human serum, living cells and biological matrices. We have utilised the long luminescence lifetime of the probe to measure ONOO À in a time-resolved manner, effectively avoiding the influence of autofluorescence in biological samples. To demonstrate the utility of the Eu(III) probe, we monitored the production of ONOO À in different cell lines, following treatment with a cold atmospheric plasma device commonly used in the clinic for skin wound treatment.

On the Liquid Chemistry of the Reactive Nitrogen Species Peroxynitrite and Nitrogen Dioxide Generated by Physical Plasmas

“…The presence of peroxynitrite [ 43 , 54 ] and peroxynitrate [ 55 ] has been reported for liquid systems under acidic conditions (pH < 4.5), as well as in alkaline conditions (pH 12) in the case of peroxynitrite [ 56 ], while information on neutral environments is missing. However, using an peroxynitrite sensitive europium probe, intracellular peroxynitrite was recently detected in human cell lines following a plasma treatment [ 57 ]. Natural sources of RNS are enzymes of the nitric oxide synthase (NOS) complex, generating ˙NO.…”

Critical Evaluation of the Interaction of Reactive Oxygen and Nitrogen Species with Blood to Inform the Clinical Translation of Nonthermal Plasma Therapy

“…Here, we used a new luminescent europium(III) probe ( Eu.1 ) to detect direct NTP-generated ONOO – in PBS, BP, and processed whole blood. This probe, designed and synthesized by some of the authors, has been previously demonstrated to rapidly and selectively detect ONOO – in a time-resolved manner, in aqueous buffers and human BP [ 28 ]. In the current study, the long luminescence lifetime of the Eu(III) probe has been utilized to eliminate any short-lived autofluorescence arising from biomolecules in the BP or processed whole blood, thereby providing high signal-to-noise and accurate measurements of ONOO – concentration.…”

“…Here, we used a new luminescent europium(III) probe ( Eu.1 , Figure 1(a) ) which we developed previously, for detection of ONOO − in vitro and in living cells [ 28 ]. For initial time points, 100 μ L of Eu.1 (100 μ M) in relevant media was treated with direct NTP and the emission intensity of Eu.1 was measured immediately.…”

“…Both RNS and H 2 O 2 , stable in PBS, were scavenged by different components of the blood. Our previously developed luminescent europium(III) probe, Eu.1 ( Figure 1(a) ), [ 28 ] was also used for precision measurement of ONOO − and revealed that NTP-generated ONOO − was present in all three liquids, for at least 30 seconds. Based on our results, we discussed the scavenging capacity of different blood components as well as the necessary considerations to choose the most optimal NTP modality for the desired clinical application.…”

Tuning the anion binding properties of lanthanide receptors to discriminate nucleoside phosphates in a sensing array

“…blood serum), 32 by using timegating methods to distinguish the lanthanide-centred emission from the short-lived autouorescence of biomolecules. [33][34][35] Time-gated (or time-resolved) luminescence measurements offer enhanced signal-to-noise and very low limits of anion detection.…”