Lanthanide‐based bulky counterions against aggregation‐caused quenching of dyes in fluorescent polymeric nanoparticles

Caterina Severi,Jaana Rosenberg,Satu Lahtinen,Tero Soukka,Andrey S Klymchenko,Andreas Reisch

doi:10.1002/agt2.130

Abstract

AbstractDye‐loaded polymeric nanoparticles (NPs) are promising bioimaging agents because of their available surface chemistry, high brightness, and tunable optical properties. However, high dye loadings can cause the aggregation‐caused quenching (ACQ) of the encapsulated fluorophores. Previously, we proposed to mitigate the ACQ inside polymeric NPs by insulating cationic dyes with bulky hydrophobic counterions. In order to implement new functionalities into dye‐loaded NPs, here, we extend the concept of bulky counterions to anionic lanthanide‐based complexes. We show that by employing Gd‐based counterions with octadecyl rhodamine B loaded NPs at 30 wt% versus polymer, the fluorescence quantum yield can be increased to 10‐fold (to 0.34). Moreover, Gd‐anion provides NPs with enhanced contrast in electron microscopy. A combination of a luminescent Eu‐based counterion with a far‐red to near‐infrared cyanine 5 dye (DiD) yields Forster resonance energy transfer NPs, where the UV‐excited Eu‐based counterion transfers energy to DiD, generating delayed fluorescence and large stokes shift of ∼340 nm. Cellular studies reveal low cytotoxicity of NPs and their capacity to internalize without detectable dye leakage, in contrast to leaky NPs with small counterions. Our findings show that the aggregation behavior of cationic dyes in the polymeric NPs can be controlled by bulky lanthanide anions, which will help in developing bright luminescent multifunctional nanomaterials.

Highlights

When dealing with charged dyes, a powerful approach is the use of bulky hydrophobic counterions (BHC),[21,30] which insulate the fluorophores effectively to avoid aggregation-caused quenching phenomenon (ACQ) and at the same time improve the dye encapsulation inside the polymeric matrix, preventing dye leaching in biological media
In the specific context of dye-loaded polymeric nanoparticles, the reasons behind preparing NPs loaded with lanthanide-based counterions are multiple: (i) lanthanides are heavy atoms routinely used as contrast agents in transmission electron microscopy (TEM), dyeloaded fluorescent nanoparticles encapsulating a lanthanide-based counterion can function as multifunctional platforms for fluorescence microscopy and TEM; (ii) europium complexes are attractive luminescent reporters due to their long-lived emission,[59,60,61] allowing background-free time-gated luminescence biosensing;[62,63,64,65] (iii) emission bands of the UV-excited europium complexes fall in the visible, making it a potential Förster resonance energy transfer (FRET) donor for the cationic fluorescent dyes coupled with such counterion, generating ultra-long Stokes shift systems
We found that NPs loaded with Gd-based counterions showed superior quantum yields and smaller diameters in respect to their counterparts based on small hydrophilic counterion

Summary

Introduction

Fluorescent nanoparticles (NPs) have emerged as powerful biosensing and bioimaging tools due to their superior optical properties in respect to organic dyes, while their available surface chemistry allows combining different functionalities in one theranostic platform.[1,2,3,4] There are several different types of fluorescent NPs, with the most famous being: quantum dots (QDs),[5,6] dye-doped silica nanoparticles[7] and carbon dots (CDs).[8,9] On the other hand, NPs assembled exclusively of organic materials have gained a large interest,[10] with systems such as aggregation-induced emission (AIE) NPs,[11,12,13] conjugated polymer NPs14,15 as well as dyeloaded lipid[16] and polymer NPs,[17,18,19] progressively becoming more popular and widespread. Lanthanide-based nanomaterials found a wide usage in theranostics applications due to their unique spectroscopic properties, such as: many narrow emission bands, long lifetimes, background-free X-ray-excited luminescence, and potentially upconversion.[56,57,58] In the specific context of dye-loaded polymeric nanoparticles, the reasons behind preparing NPs loaded with lanthanide-based counterions are multiple: (i) lanthanides are heavy atoms routinely used as contrast agents in transmission electron microscopy (TEM), dyeloaded fluorescent nanoparticles encapsulating a lanthanide-based counterion can function as multifunctional platforms for fluorescence microscopy and TEM; (ii) europium complexes are attractive luminescent reporters due to their long-lived emission,[59,60,61] allowing background-free time-gated luminescence biosensing;[62,63,64,65] (iii) emission bands of the UV-excited europium complexes fall in the visible, making it a potential Förster resonance energy transfer (FRET) donor for the cationic fluorescent dyes coupled with such counterion, generating ultra-long Stokes shift systems We tested these lanthanide-based counterions by formulating NPs loaded with two cationic dyes: an octadecyl ester of rhodamine B (R18) and a dioctadecyl derivative of cyanine 5 (DiD) (Figure 1A). The NPs loaded with dye paired with the lanthanide anion displayed minimal cytotoxicity at concentrations habitually used for cellular imaging

Preparation of NPs based on lanthanide counterions

Single-particle measurements

Findings

Conclusion