Affimer-Based Europium Chelates Allow Sensitive Optical Biosensing in a Range of Human Disease Biomarkers

Abstract

The protein biomarker measurement has been well-established using ELISA (enzyme-linked immunosorbent assay), which offers good sensitivity and specificity, but remains slow and expensive. Certain clinical conditions, where rapid measurement or immediate confirmation of a biomarker is paramount for treatment, necessitate more rapid analysis. Biosensors offer the prospect of reagent-less, processing-free measurements at the patient’s bedside. Here, we report a platform for biosensing based on chelated Eu3+ against a range of proteins including biomarkers of cardiac injury (human myoglobin), stroke (glial fibrillary acidic protein (GFAP)), inflammation (C-reactive protein (CRP)) and colorectal cancer (carcinoembryonic antigen (CEA)). The Eu3+ ions are chelated by modified synthetic binding proteins (Affimers), which offer an alternative targeting strategy to existing antibodies. The fluorescence characteristics of the Eu3+ complex with modified Affimers against human myoglobin, GFAP, CRP and CEA were measured in human serum using λex = 395 nm, λem = 590 and 615 nm. The Eu3+-Affimer based complex allowed sensitive detection of human myoglobin, GFAP, CRP and CEA proteins as low as 100 fM in (100-fold) diluted human serum samples. The unique dependence on Eu3+ fluorescence in the visible region (590 and 615 nm) was exploited in this study to allow rapid measurement of the analyte concentration, with measurements in 2 to 3 min. These data demonstrate that the Affimer based Eu3+ complexes can function as nanobiosensors with potential analytical and diagnostic applications.

Recent Progress in Biosensors for Detection of Tumor Biomarkers

“…To further improve biosensors, Al-Enezi et al developed a novel technique using Affimer-based Eu 3+ complexes as nanobiosensors for the optical biosensing of CEA. This method has high sensitivity, with a detection limit below 100 fM, especially in clinical trials as a targeting strategy [ 42 ].…”

Investigation of Peptides for Molecular Recognition of C-Reactive Protein–Theoretical and Experimental Studies

“…Antibodies are currently the most frequently used CRP receptors. 24,29,30 In the literature, there are also examples of CRP determination using bioreceptors such as aptamers, 31−35 bacteriophages, 36 affimers, 37 peptides, 38,39 or nanobodies. 40 CRP assays are often performed using biological methods such as ELISA, 41 test-based lateral flow immunoassays, and other immunoassays.…”

“…Antibodies are currently the most frequently used CRP receptors. ,, In the literature, there are also examples of CRP determination using bioreceptors such as aptamers, – bacteriophages, affimers, peptides, , or nanobodies …”

“…A solution might be characterization of the molecular interactions of CRP with receptors using various physicochemical techniques. Some examples are 2D DNA origami nanostructures for quantitative single-molecule biosensing, SERS, fluorescence microscopy, atomic force microscopy, biolayer interferometry, or SPR ,, and electrochemical methods. ,,– Not all of these are easy to perform, but they can give deeper analysis and insights into understanding protein–peptide interactions.…”

Layered Gadolinium-Europium-Terbium Hydroxides Sensitised with 4-Sulfobenzoate as All Solid-State Luminescent Thermometers

“…Primarily, such materials include sensors providing a negative [25,26] or positive [27,28] luminescence response to the detected molecules. Thus, a biosensor for the detection and measurement of a range of protein biomarkers in human serum was created on the basis of an Eu 3+ -complex, with a detection limit of <100 fM [29]. A terbium(III) complex with 1,10phenanthroline was demonstrated to be capable of detecting ascorbic acid with a detection limit of 7.4 × 10 −5 mol L −1 [30].…”