Fluorescence Resonance Energy Transfer Technique Research Articles

Harnessing fluorescence resonance energy transfer for improved solar still performance with zinc oxide nanoparticles and activated carbon

The Single Slope Corrugated Wick type Solar Distillers (SWS) have attracted significant research funding owing to their simple preparation process and excellent durability against heat and moisture. However, there is a recognized need to enhance their solar energy conversion efficiency. The utilization of ZnO nanoparticles (Z) in SWS is limited by their light absorption capacity, resulting in suboptimal thermal energy conversion and solar energy absorption. In this investigation, we propose an innovative method known as Fluorescence Resonance Energy Transfer (FRET), combining crystallized Z with activated carbon (AC) derived from areca nut shells. This novel approach substantially boosts the thermal efficiency of solar energy utilization in SWS by facilitating efficient energy transfer from ZAC to water.The FRET technique enables the effective transfer of energy from ZAC to water and is comprehensively analyzed. By utilizing Z as donors and AC as acceptors, the photoluminescence emitted by ZAC can be absorbed by both components, thereby increasing the carrier population as of Corrugated Wick-type towards the condenser face within SWS. The evaporation rate from water to the condensing surface is significantly higher in SWS with ZAC compared to conventional wick-type solar stills (CWS), attributed to the efficient heat transfer facilitated through multi-dimensional heterojunctions within ZAC. The energy and charge transfer processes are optimized for both summer and winter operating conditions. The ZAC-based SWS exhibits exceptional resilience in various ambient conditions, resulting in a substantial 33.989 % enhancement in distillation efficiency during summer and a notable 31.199 % improvement in winter. The incorporation of ZAC elevates the production of distillate water to 6.325 liters/m2 per day in summer and 5.840 liters/m2 per day in winter (24 h). The energy distribution among particle components is extensively scrutinized using the FRET mechanism.

A Sustainable Retinal Drug Co-Delivery for Boosting Therapeutic Efficacy in wAMD: Unveiling Multifaceted Evidence and Synergistic Mechanisms.

Sustainable retinal codelivery poses significant challenges technically, although it is imperative for synergistic treatment of wet age-related macular degeneration (wAMD). Here, a microemulsion-doped hydrogel (Bor/PT-M@TRG) is engineered as an intravitreal depot composing of temperature-responsive hydrogel (TRG) and borneol-decorated paeoniflorin (PF) & tetramethylpyrazine (TMP)-coloaded microemulsions (Bor/PT-M). Bor/PT-M@TRG, functioning as the "ammunition depot", resides in the vitreous and continuously releases Bor/PT-M as the therapeutic "bullet", enabling deep penetration into the retina for 21 days. A single intravitreal injection of Bor/PT-M@TRG yields substantial reductions in choroidal neovascularization (CNV, a hallmark feature of wAMD) progression and mitigates oxidative stress-induced damage in vivo. Combinational PF&TMP regulates the "reactive oxygen species/nuclear factor erythroid-2-related factor 2/heme oxygenase-1" pathway and blocks the "hypoxia inducible factor-1α/vascular endothelial growth factor" signaling in retina, synergistically cutting off the loop of CNV formation. Utilizing fluorescence resonance energy transfer and liquid chromatography-mass spectrometry techniques, they present compelling multifaceted evidence of sustainable retinal codelivery spanning formulations, ARPE-19 cells, in vivo eye balls, and ex vivo section/retina-choroid complex cell levels. Such codelivery approach is elucidated as the key driving force behind the exceptional therapeutic outcomes of Bor/PT-M@TRG. These findings highlight the significance of sustainable retinal drug codelivery and rational combination for effective treatment of wAMD.

Association of CC-chemokine ligand-2 gene polymorphisms with leprosy reactions

BackgroundC–C motif chemokine ligand 2, a gene that codes for a protein involved in inflammation. Certain SNPs in the CCL2 gene have been studied for their potential associations with susceptibility to various diseases. These SNPs may affect the production and function of the CCL2 protein, which is involved in the recruitment of immune cells to the site of inflammation. Variations in CCL2 may influence the immune response to Mycobacterium leprae infection. ObjectiveTo investigate the association of the C–C motif chemokine ligand-2 single nucleotide polymorphisms with leprosy. MethodsCCL2 single nucleotide polymorphisms were analyzed in a total of 975 leprosy patients and 357 healthy controls. Of those, 577 leprosy and 288 healthy controls were analyzed by PCR-RFLP for CCL2 -2518 A>G, 535 leprosy and 290 controls for CCL2 -362 G>C, 295 leprosy and 240 controls for CCL2 -2134 T>G, 325 leprosy and 288 controls for CCL2 -1549 A>T SNPs by melting curve analysis using hybridization probe chemistry and detection by fluorescence resonance energy transfer (FRET) technique in Realtime PCR. The levels of CCL2, IL-12p70, IFN-γ, TNF-α, and TGF-β were estimated in sera samples and correlated with CCL2 genotypes. ResultsThe frequency of the GCT (-2518 A>G, -362 G>C, -2134 T>G) haplotype is observed to be higher in leprosy patients compared to healthy controls (P = 0.04). There was no significant difference observed in genotypic frequencies between leprosy patients and healthy controls {(-2518A>G, p = 0.53), (-362 G>C, p = 0.01), (-2134 T>G, p = 0.10)}. G allele at the -2134 site is predominant in leprosy (borderline) without any reaction (8 %) compared to borderline patients with RR reactions (2.1 %) (P = 0.03). GG genotype (p = 0.008) and G allele at -2518 (p = 0.030) of the CCL 2 gene were found to be associated with patients with ENL reaction. An elevated level of serum CCL2 was observed in leprosy patients with the -2518 AA and AG genotypes (p = 0.0001). ConclusionsG allele and GG genotype at the CCL2 -2518 site are associated with a risk of ENL reactions.

Cellulose nanocrystals from celery stalk as quercetin scaffolds: A novel perspective of human holo-transferrin adsorption and digestion behaviours.

In this study, cellulose nanocrystals (CNCs) were synthesized from celery stalks to be used as the platform for quercetin delivery. Additionally, CNCs and CNCs-quercetin were characterized using the results of scanning electron microscope (SEM), transmission electron microscopy (TEM), X-ray diffraction (XRD), Fourier transform infrared (FTIR) spectroscopy, and zeta potential, while their interactions with human holo-transferrin (HTF) were also investigated. We examined their interaction under physiological conditions through the exertion of fluorescence, resonance light scattering, synchronized fluorescence spectroscopy, circular dichroism, three-dimensional fluorescence spectroscopy, and fluorescence resonance energy transfer techniques. The data from SEM and TEM exhibited the spherical shape of CNCs and CNCs-quercetin and also, a decrease was detected in the size of quercetin-loaded CNCs from 676 to 473 nm that indicated the intensified water solubility of quercetin. The success of cellulose acid hydrolysis was confirmed based on the XRD results. Apparently, the crystalline index of CNCs-quercetin was reduced by the interaction of CNCs with quercetin, which also resulted in the appearance of functional groups, as shown by FTIR. The interaction of CNCs-quercetin with HTF was also demonstrated by the induced quenching in the intensity of HTF fluorescence emission and Stern-Volmer data represent the occurrence of static quenching. Overall, the effectiveness of CNCs as quercetin vehicles suggests its potential suitability for dietary supplements and pharmaceutical products.

Heterodimerization of Human UDP-Glucuronosyltransferase 1A9 and UDP-Glucuronosyltransferase 2B7 Alters Their Glucuronidation Activities.

Human UDP-glucuronosyltransferases (UGTs) play a pivotal role as prominent phase II metabolic enzymes, mediating the glucuronidation of both endobiotics and xenobiotics. Dimerization greatly modulates the enzymatic activities of UGTs. In this study, we examined the influence of three mutations (H35A, H268Y, and N68A/N315A) and four truncations (signal peptide, single transmembrane helix, cytosolic tail, and di-lysine motif) in UGT2B7 on its heterodimerization with wild-type UGT1A9, using a Bac-to-Bac expression system. We employed quantitative fluorescence resonance energy transfer (FRET) techniques and co-immunoprecipitation assays to evaluate the formation of heterodimers between UGT1A9 and UGT2B7 allozymes. Furthermore, we evaluated the glucuronidation activities of the heterodimers using zidovudine and propofol as substrates for UGT2B7 and UGT1A9, respectively. Our findings revealed that the histidine residue at codon 35 was involved in the dimeric interaction, as evidenced by the FRET efficiencies and catalytic activities. Interestingly, the signal peptide and single transmembrane helix domain of UGT2B7 had no impact on the protein-protein interaction. These results provide valuable insights for a comprehensive understanding of UGT1A9/UGT2B7 heterodimer formation and its association with glucuronidation activity. SIGNIFICANCE STATEMENT: Our findings revealed that the H35A mutation in UGT2B7 affected the affinity of protein-protein interaction, leading to discernable variations in fluorescence resonance energy transfer efficiencies and catalytic activity. Furthermore, the signal peptide and single transmembrane helix domain of UGT2B7 did not influence heterodimer formation. These results provide valuable insights into the combined effects of polymorphisms and protein-protein interactions on the catalytic activity of UGT1A9 and UGT2B7, enhancing our understanding of UGT dimerization and its impact on metabolite formation.

Carbon quantum dots for efficient delivery of curcumin in live cell

Carbon quantum dots (CQD) are a novel class of nanomaterials that has significant importance for applications in bioimaging, drug loading and delivery. Their easy preparation, tunable optoelectronic property, low toxicity, excellent biocompatibility, superior photostability, and exceptional water solubility contribute to their tremendous potentials for various applications. In this study, we synthesized carbon quantum dots using a greener and cost-efficient top-down reflux methods, and characterized the CQDs by various spectroscopy techniques. Furthermore, we investigated the potential application of CQDs as carrier of antitumor drug curcumin (Cur) in vitro by investigating the drug loading and release mechanism of the CQDs. Solubility of hydrophobic drug curcumin is increased when it forms complex with CQD, and hence increases the potential for its application. Moreover, the pH responsive drug release mechanism of the curcumin loaded CQDs is found to follow anomalous diffusion as per Korsmeyer-Peppas model. Additionally, we explored the in vivo bioimaging application of the CQDs, and used fluorescence resonance energy transfer (FRET) technique using CQDs (as donor) and curcumin (as acceptor) to understand the heterogeneity of drug loading in live Escherichia coli (E. coli) cells. Our findings provide a detailed insight into the application of CQDs as probes in bioimaging and FRET, and as well as their potential for targeted curcumin delivery in live cells.

Dynamics of nuclear export of pre-ribosomal subunits revealed by high-speed single-molecule microscopy in live cells

We present a study on the nuclear export efficiency and time of pre-ribosomal subunits in live mammalian cells, using high-speed single-molecule tracking and single-molecule fluorescence resonance energy transfer techniques. Our findings reveal that pre-ribosomal particles exhibit significantly higher nuclear export efficiency compared to other large cargos like mRNAs, with around two-thirds of interactions between the pre-60S or pre-40S and the nuclear pore complexes (NPCs) resulting in successful export to the cytoplasm. We also demonstrate that nuclear transport receptor (NTR) chromosomal maintenance 1 (CRM1) plays a crucial role in nuclear export efficiency, with pre-60S and pre-40S particle export efficiency decreasing by 11-17-fold when CRM1 is inhibited. Our results suggest that multiple copies of CRM1 work cooperatively to chaperone pre-ribosomal subunits through the NPC, thus increasing export efficiency and decreasing export time. Significantly, this cooperative NTR mechanism extends beyond pre-ribosomal subunits, as evidenced by the enhanced nucleocytoplasmic transport of proteins.

Parallel DNA G-Quadruplex Induced and Stabilized by Curaxin CBL0137

G-quadruplex (G4) is one of the higher-order DNA structures in guanine-rich sequences which are widely distributed across the genome. Due to their presence in oncogenic promoters and telomeres, G4 DNA structures become the novel targets in anticancer drug designs. Curaxin CBL0137, as an important candidate anticancer drug, can effectively inhibit the growth of multiple cancers. Although there is evidence that anticancer activity of curaxin is associated with its ability to bind DNA and to change the DNA topology, its therapeutic target and the underlying anti-cancer mechanism are still unclear. Here we show, for the first time, that curaxin CBL0137 induces G4 folding from anti-parallel to parallel structures, by single-molecule fluorescence resonance energy transfer technique. More importantly, we find that curaxin CBL0137 promotes G4 folding as well as stabilizes the folded G4 structures with long loops, giving a novel insight into effects of curaxin CBL0137 on DNA structures. Our work provides new ideas for the therapeutic mechanism of curaxin CBL0137 and for designs of new G4-targeting anticancer drugs.

Green Nanoformulations of Polyvinylpyrrolidone-Capped Metal Nanoparticles: A Study at the Hybrid Interface with Biomimetic Cell Membranes and In Vitro Cell Models.

Noble metal nanoparticles (NP) with intrinsic antiangiogenic, antibacterial, and anti-inflammatory properties have great potential as potent chemotherapeutics, due to their unique features, including plasmonic properties for application in photothermal therapy, and their capability to slow down the migration/invasion speed of cancer cells and then suppress metastasis. In this work, gold (Au), silver (Ag), and palladium (Pd) NP were synthesized by a green redox chemistry method with the reduction of the metal salt precursor with glucose in the presence of polyvinylpyrrolidone (PVP) as stabilizing and capping agent. The physicochemical properties of the PVP-capped NP were investigated by UV-visible (UV-vis) and attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopies, dynamic light scattering (DLS), and atomic force microscopy (AFM), to scrutinize the optical features and the interface between the metal surface and the capping polymer, the hydrodynamic size, and the morphology, respectively. Biophysical studies with model cell membranes were carried out by using laser scanning confocal microscopy (LSM) with fluorescence recovery after photobleaching (FRAP) and fluorescence resonance energy transfer (FRET) techniques. To this purpose, artificial cell membranes of supported lipid bilayers (SLBs) made with 1-palmitoyl-2-oleoyl-sn-glycerol-3-phosphocholine (POPC) dye-labeled with 7-nitro-2-1,3-benzoxadiazol-4-yl (NBD, FRET donor) and/or lissamine rhodamine B sulfonyl (Rh, FRET acceptor) were prepared. Proof-of-work in vitro cellular experiments were carried out with prostate cancer cells (PC-3 line) in terms of cytotoxicity, cell migration (wound scratch assay), NP cellular uptake, and cytoskeleton actin perturbation.

A multiparametric fluorescent visualization approach for detecting drug resistance in living cancer cells

Drug resistance is a worldwide health care crisis which impedes disease treatment and increases financial burden, especially for its multifactorial nature and high complexity. Herein, we developed a multiparametric approach to visualize and detect drug resistance in living cancer cells, through the combination of DNA-templated covalent protein labeling strategy and fluorescent resonance energy transfer technique. Gefitinib resistance in non-small cell lung cancer caused by mesenchymal-epidermal transition factor (Met) overexpression and hyperactivation was investigated as a proof-of-concept. Unlike the traditional single-factor investigation, the proposed approach evaluated the contribution of three important parameters towards the resistance, including the changes of Met expression level, the homodimerization of Met with itself and the heterodimerization of Met with epidermal growth factor receptor (EGFR). A multiple regression model based on these three parameters was tentatively established for evaluation of the resistance level of laboratory-developed resistant cells and evaluation of the resistance level of patient-derived cells. Such an approach facilitates a quick identification of a drug resistance, to evaluate not only the resistance level but also the resistance mechanism.

A FRET Based Composite Sensing Material Based on UCNPs and Rhodamine Derivative for Highly Sensitive and Selective Detection of Fe3.

The existence of excessive concentration of iron ion (Fe3+) in water will do harm to the environment and biology. Presently, sensitive and selective determination of Fe3+ directly in real environment samples is still a challenging job because of the high complexity of the sample matrix. In this work, we reported a new sensor system for Fe3+ based on fluorescence resonance energy transfer (FRET) from upconversion nanoparticles (UCNPs) to Rhodamine derivative probe (RhB). The NaYF4: Yb, Er@SiO2@P(NIPAM-co-RhB) nanocomposites was constructed, in which PNIPAm was used as the probe carrier. The nanocomposites can not only be excited by infrared light to avoid the interference of background light in the Fe3+ detection process, but also enhance the detection signal output through temperature control. Under the optimum conditions, the RSD (Relative standard deviation) of actual sample measurements ranges was from 1.95% to 4.96%, with the recovery rate from 97.4% to 103.3%, which showed high reliability for Fe3+ detection. This work could be extended to sensing other target ions or molecules and may promote the widespread use of FRET technique.

Single-Molecule Study of DNAzyme Reveals Its Intrinsic Conformational Dynamics.

DNAzyme is a class of DNA molecules that can perform catalytic functions with high selectivity towards specific metal ions. Due to its potential applications for biosensors and medical therapeutics, DNAzyme has been extensively studied to characterize the relationships between its biochemical properties and functions. Similar to protein enzymes and ribozymes, DNAzymes have been found to undergo conformational changes in a metal-ion-dependent manner for catalysis. Despite the important role the conformation plays in the catalysis process, such structural and dynamic information might not be revealed by conventional approaches. Here, by using the single-molecule fluorescence resonance energy transfer (smFRET) technique, we were able to investigate the detailed conformational dynamics of a uranyl-specific DNAzyme 39E. We observed conformation switches of 39E to a folded state with the addition of Mg2+ and to an extended state with the addition of UO22+. Furthermore, 39E can switch to a more compact configuration with or without divalent metal ions. Our findings reveal that 39E can undergo conformational changes spontaneously between different configurations.

In situ self-assembly of amphiphilic dextran micelles and superparamagnetic iron oxide nanoparticle-loading as magnetic resonance imaging contrast agents.

Polymeric micelles have long been considered as promising nanocarrier for hydrophobic drugs and imaging probes, due to their nanoscale particle size, biocompatibility and ability to loading reasonable amount of cargoes. Herein, a facile method for dextran micelles preparation was developed and their performance as carriers of superparamagnetic iron oxide (SPIO) nanocrystals was evaluated. Amphiphilic dextran (Dex-g-OA) was synthesized via the Schiff base reactions between oxidized dextran and oleylamine, and self-assembled in situ into nano-size micelles in the reaction systems. The self-assembling behaviors of the amphiphilic dextran were identified using fluorescence resonance energy transfer technique by detection the energy transfer signal between the fluorophore pairs, Cy5 and Cy5.5. Hydrophobic SPIO nanoparticles (Fe3O4 NPs) were successfully loaded into the dextran micelles via the in situ self-assembly process, leading to a series of Fe3O4 NPs-loaded micelle nanocomposites (Fe3O4@Dex-g-OA) with good biocompatibility, superparamagnetism and strongly enhanced T 2 relaxivity. At the magnetic field of 0.5 T, the Fe3O4@Dex-g-OA nanocomposite with particle size of 116.2 ± 53.7 nm presented a higher T 2 relaxivity of 327.9 ·s-1. The prepared magnetic nanocomposites hold the promise to be used as contrast agents in magnetic resonance imaging.

FLIM-FRET Investigation of Heterogeneous Huntingtin Aggregation in HeLa Cells.

Fluorescence lifetime imaging microscopy-fluorescence resonance energy transfer (FLIM-FRET) technique is a novel tool to study protein-protein interaction combining FRET detection, lifetime measurements, and high-resolution imaging. This chapter describes a FLIM-FRET protocol for the detection and quantification of heterogeneous huntingtin aggregates in HeLa cells co-transfected with huntingtin variants containing polyglutamine tracts of different lengths.

Investigating Interactions Between Histone Modifying Enzymes and Transcription Factors <em>in vivo</em> by Fluorescence Resonance Energy Transfer

Epigenetic regulation of gene expression is commonly affected by histone modifying enzymes (HMEs) that generate heterochromatic or euchromatic histone marks for transcriptional repression or activation, respectively. HMEs are recruited to their target chromatin by transcription factors (TFs). Thus, detecting and characterizing direct interactions between HMEs and TFs are critical for understanding their function and specificity better. These studies would be more biologically relevant if performed in vivo within living tissues. Here, a protocol is described for visualizing interactions in plant leaves between a plant histone deubiquitinase and a plant transcription factor using fluorescence resonance energy transfer (FRET), which allows the detection of complexes between protein molecules that are within <10 nm from each other. Two variations of the FRET technique are presented: SE-FRET (sensitized emission) and AB-FRET (acceptor bleaching), in which the energy is transferred non-radiatively from the donor to the acceptor or emitted radiatively by the donor upon photobleaching of the acceptor. Both SE-FRET and AB-FRET approaches can be adapted easily to discover other interactions between other proteins in planta.

Investigating Interactions Between Histone Modifying Enzymes and Transcription Factors in vivo by Fluorescence Resonance Energy Transfer.

Epigenetic regulation of gene expression is commonly affected by histone modifying enzymes (HMEs) that generate heterochromatic or euchromatic histone marks for transcriptional repression or activation, respectively. HMEs are recruited to their target chromatin by transcription factors (TFs). Thus, detecting and characterizing direct interactions between HMEs and TFs are critical for understanding their function and specificity better. These studies would be more biologically relevant if performed in vivo within living tissues. Here, a protocol is described for visualizing interactions in plant leaves between a plant histone deubiquitinase and a plant transcription factor using fluorescence resonance energy transfer (FRET), which allows the detection of complexes between protein molecules that are within <10 nm from each other. Two variations of the FRET technique are presented: SE-FRET (sensitized emission) and AB-FRET (acceptor bleaching), in which the energy is transferred non-radiatively from the donor to the acceptor or emitted radiatively by the donor upon photobleaching of the acceptor. Both SE-FRET and AB-FRET approaches can be adapted easily to discover other interactions between other proteins in planta.

Measuring Endocannabinoid System Interaction with Biomembranes.

Understanding the correct interaction among the different components of the endocannabinoid (eCB) system is fundamental for a proper assessment of the function of eCBs as signaling molecules. The knowledge of how the membrane environment modulates the intracellular trafficking of the eCB system and its interacting proteins holds a huge potential in unraveling new mechanisms of its modulation. This chapter deals with the application of fluorescence resonance energy transfer technique to measure the binding affinity of eCB proteins to model membranes (i.e., large unilamellar vesicles, LUVs). In particular, we describe in detail the paradigmatic example of the interaction of rat recombinant fatty acid amide hydrolase with LUVs constituted of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine.

Progress of target determination and mechanism of bioactive components of traditional Chinese medicine

The pharmacodynamic substances of traditional Chinese medicine(TCM) are the basis for the research of TCM and the development of innovative drugs. However, the lack of clarity of targets and molecular mechanisms is the bottleneck problem that restricts the research of pharmacodynamic substances of TCM. Bioactive components are the material basis of the efficacy of TCM, which exert activity by regulating the corresponding targets. Therefore, it is very important to identify the targets of the bioactive components to elucidate the pharmacological mechanism of TCM. Proteins are the most important drug targets, and study of the interaction between the proteins and bioactive components of TCM plays a key role in the development of pharmacological mechanism of TCM. In recent years, the main techniques for detecting the interaction between the bioactive components and proteins include surface plasmon resonance, fluorescence resonance energy transfer, bio-layer interference, molecular docking, proteome chip, target fishing, target mutant, and protein crystallization techniques, etc. This review summarized the biological target detection techniques and their applications in locating the targets of the bioactive components in TCM in the last decade, and this paper will provide useful strategies to elucidate the pharmacological mechanisms of TCM.

Fluorescence interactions of a novel chalcone derivative with membrane model systems and human serum albumin

A novel chalcone derivative (4-(5-(pyridin-2-yl)-4,5-dihydro-1H-pyrazol-3-yl) phenol (PDP) was synthesized, characterized and investigated for its potential as a fluorescent probe. The structure of the synthesized molecule was determined by FTIR, 1H NMR, 13C NMR and LC-MS/MS. The interactions of PDP with fluorescent dyes in aqueous SDS environment and HSA were studied by using fluorescence resonance energy transfer (FRET) technique and steady-state and time-resolved fluorescence spectroscopy techniques. In addition, the cytotoxic effects of PDP against various cell lines (MCF -7, HT −29, and 3 T3-L1) as well as their corresponding healthy cell lines were tested by MTT assay and visualization of FRET efficiency of PDP in vitro was monitored by confocal microscopy. MTT assay showed that PDP has no significant cytotoxic effect on HT −29 cancer cells and moderate cytotoxicity on MCF -7 and 3 T3-L1 cells even at a concentration of 250 μM. Combining confocal microscopes with the FRET technique showed that PDP significantly stained the cytoplasm of MCF -7 cell lines. These results suggest that PDP could be used in fluorescence microscopy for cell staining.

ADAMTS13 levels in a plasma-derived FVIII concentrate: A potential therapeutic option for patients with congenital thrombotic thrombocytopenic purpura

IntroductionTreatment of congenital thrombotic thrombocytopenic purpura (cTTP), a disease characterized by the congenital deficiency of ADAMTS13, remains a challenge as there are no specific treatments available yet, other than therapy based on the use of fresh frozen plasma (FFP). Since cTTP is caused by low levels of ADAMTS13 protein, commercially available coagulation factor concentrates have been considered as potential ADAMTS13 source in place of FFP. The study aimed to validate the therapeutic potential of a plasma-derived factor VIII (FVIII) product as a source of ADAMTS13. MethodsThe quantitation of ADAMTS13 activity levels in eight lots of a plasma-derived FVIII product, (Koāte®) was carried out with three different methodologies: a Fluorescence Resonance Energy Transfer (FRET) assay, a chemiluminescence assay, and a chromogenic ELISA. ADAMTS13 protein antigen levels were measured by the FRET technique as well. In addition, von Willebrand factor (VWF) activity (VWF ristocetin cofactor, VWF:RCo, and VWF collagen binding, VWF:CB) and antigen (VWF:Ag) were measured using chemiluminescence assays. Qualification protocols were applied to the methods used. ResultsThe results showed high levels of ADAMTS13 in all eight Koāte® lots analyzed, with antigen and activity levels respectively of 10.72 IU/ml ± 3.94 and 5.62 IU/ml ± 1.39. Despite the significant content of ADAMTS13, VWF integrity seems not to be affected (0.81 ± 0.15 VWF:RCo/VWF:Ag and 0.75 ± 0.15 VWF:CB/VWF:Ag ratios). ConclusionsThese findings suggest that Koāte® could be a potential candidate for the treatment of cTTP, warranting evaluation in a clinical trial.