Can the Dose of Belimumab be Reduced in Patients with Systemic Lupus Erythematosus?

OBJECTIVES: The aims of this study were to investigate the prevalence of dose reduction in patients with SLE treated with belimumab (BEL) in Spain, analyze treatment modalities, and determine impact on control of disease activity. METHODS: Retrospective longitudinal and multicentre study of SLE patients treated with BEL. Data on disease activity, treatments and outcomes were recorded before and after reduction (6-12 months), and they were compared. RESULTS: A total of 324 patients were included. The dose was reduced in 29 patients (8.9%). The dosing interval was increased in 9 patients receiving subcutaneous BEL and in 6 patients receiving intravenous BEL. The dose per administration was reduced in 16 patients.Pre-reduction status was remission (2021 DORIS) in 15/26 patients (57.7%) and LLDAS in 23/26 patients (88.5%). After reduction, 2/24 patients (8.3%) and 3/22 patients (13.6%) lost remission at 6 months and 12 months, respectively (not statistically significant [NS]). As for LLDAS, 2/23 patients (8.7%) and 2/21 patients (9.5%) lost their status at 6 and 12 months, respectively (NS). Significantly fewer patients were taking glucocorticoids (GCs) at their 12-month visit, although the median dose of GCs was higher at the 12-month visit (5 [0.62-8.75] vs 2.5 [0-5] at baseline). CONCLUSION: Doses of BEL can be reduced with no relevant changes in disease activity-at least in the short term-in a significant percentage of patients, and most maintain the reduced dose. However, increased clinical or serologic activity may be observed in some patients. Consequently, tighter post-reduction follow-up is advisable.

Is there an association between periodontal disease and infertility? A systematic review.

BACKGROUND: Today, idiopathic infertility is becoming more frequent, affecting more than 186 million people in the world. The presence of comorbidities makes patient management complex, requiring individualized infertility treatment. Periodontal diseases could contribute negatively to the management of infertility, increasing inflammation, oxidative stress and insulin resistance, and contributing negatively to the development and progression of comorbidities associated with these two entities. The aim of this systematic review is to explore whether there is an association between periodontal diseases and male and female infertility and deepen into the possible mechanisms underlying this association. MATERIAL AND METHODS: The studies analyzed in this research included a total of 4871 patients (732 men and 4139 women), were original studies with high quality, mostly with a control group. Authors who have investigated idiopathic infertility suggest that infertility is associated with diseases that present low-grade chronic inflammation, oxidative stress and insulin resistance (such as obesity, type 2 diabetes and polycystic ovary syndrome), which are in turn related to periodontal diseases. RESULTS: A higher prevalence of periodontal disease was found in patients with infertility compared with controls. Periodontal diseases could initially be mediated by a local and systemic proinflammatory environment, which favors a pro-oxidant state, leading to oxidative stress and, finally, irreversible destruction of the periodontal tissue. Insulin resistance, oxidative stress and inflammation are present in the pathologies associated with an increase in the prevalence and severity of periodontal diseases (such as obesity, type 2 diabetes and polycystic ovary syndrome). Therefore, IR, low-grade chronic inflammation and the oxidative stress could be the pathophysiological mechanisms linking idiopathic infertility and periodontal diseases. CONCLUSIONS: Studies suggest an association between infertility and periodontitis. Future researches are necessary to find causality factors. Studying the patient in a global and multidisciplinary way could help in the management and treatment of idiopathic infertility.

Role of Brazilian bats in the epidemiological cycle of potentially zoonotic pathogens.

Bats (Chiroptera) are flying mammals of great biodiversity and habits. These characteristics contribute for them being natural reservoirs and part of the epidemiological cycle of several potentially zoonotic pathogens, such as viruses, protozoa, fungi and bacteria. Brazil hosts approximately 15% of the world's bat diversity, with 181 distinct species, 68 genera and 9 families. About 60% of infectious diseases in humans are of zoonotic origin and, in the last decades, the detection of zoonotic pathogens in bats and their environment has been reported, such as Rabies virus (RABV) and Histoplasma capsulatum. Thus, the aim of this work was to review the reports of zoonotic pathogens associated with bats in Brazil in the past ten years. We reviewed the main pathogenic microorganisms described and the species of bats most frequently involved in the epidemiological cycles of these zoonotic agents. The obtained data show an upward trend in the detection of zoonotic pathogens in Brazilian bats, such as RABV, Bartonella sp., Histoplasma capsulatum and Leishmania spp., with emphasis on the bat species Artibeus lituratus, Carollia perspicillata, Desmodus rotundus and Molossus molossus. These findings highlight the importance of monitoring bat-associated microrganisms to early identify pathogens that may threaten bat populations, including potentially zoonotic microrganisms, emphasizing the importance of the One Health approach to prevent and mitigate the risks of the emergence of zoonotic diseases.

Comparison of three fluids for calibration of the new Periotron® 8010.

BACKGROUND: The aim of the present study was to calibrate the Periotron® model 8010 with volumes of three different fluids (distilled water, serum, and saliva) and to identify which of the three is the most reliable, feasible, and reproducible for routine calibration. MATERIAL AND METHODS: A total of 450 samples of Periopaper® were divided into three groups (150 each per group): distilled water, serum matrix and saliva. A calibration curve was run with 0.25, 0.50, 0.75, 1.00 and 1.25 µl of each of the fluids, and the results were determined in Periotron units (PU). Statistical analysis was performed with one-way ANOVA followed by Bonferroni's post hoc test and a linear equation. RESULTS: Distilled water presented the lowest levels of PU at all volumes, while serum showed the highest levels at high volumes. Linear regression equations rendered similar slopes for saliva and distilled water, while serum was statistically different. Saliva presented a reproduction percentage of 99.7%, which indicated better accuracy and precision than serum and distilled water. CONCLUSIONS: Saliva is more reliable and accurate than water or serum for the purpose of calibration of the Periotron® model 8010, though it shares drawbacks with serum. Distilled water is more easily available and does not require any additional procedure, in addition to producing a similar slope to saliva and a smaller deviation from the media than serum.

Salivary biomarkers present in patients with periodontitis without clinical distinction: findings from a meta-analysis.

BACKGROUND: A new classification for periodontitis has been adopted in clinical practice. However, there are still discussions regarding this new classification and difficulties in its adoption, both by professionals and researchers. Thus, this study aimed to evaluate which salivary biomarkers are present in periodontitis, following the new classification of periodontal diseases through meta-analysis. MATERIAL AND METHODS: A literature search was carried out in the scientific databases: PubMed, Scielo and Google scholar to select studies. The selection of studies was followed by two authors upon reading of the title, abstract and full text. The necessary data were collected and statistical analyses were performed using the Review Manager statistical software version 5.4, with calculation of Mean Difference, heterogeneity (I²) and funnel plot with P < 0.05. RESULTS: After following the selection criteria, 9 articles were selected for comparison. The studies address the presence of biomarkers in the saliva of patients with periodontitis and their possible use in the monitoring and diagnosis of the disease. For the meta-analytic comparison, a sample size of 1,983 individuals was used. Statistical analyses showed that nitric oxide, IL-6, IL-1B and osteoprotegerin are substances that are significantly present in patients with periodontitis (P < 0.05). CONCLUSIONS: IL-6, nitric oxide, IL-1B, TNF-α and osteoprotegerin are among the most present biomarkers in patients with periodontitis, and may be used in the future as a monitoring of periodontal disease. The present study also revealed that there was no statistically significant difference in the concentration of these biomarkers for clinical distinction from periodontitis.

EEF1A1 transcription cofactor gene polymorphism is associated with muscle gene expression and residual feed intake in Nelore cattle.

Cis-acting effects of noncoding variants on gene expression and regulatory molecules constitute a significant factor for phenotypic variation in complex traits. To provide new insights into the impacts of single-nucleotide polymorphisms (SNPs) on transcription factors (TFs) and transcription cofactors (TcoF) coding genes, we carried out a multi-omic analysis to identify cis-regulatory effects of SNPs on these genes' expression in muscle and describe their association with feed efficiency-related traits in Nelore cattle. As a result, we identified one SNP, the rs137256008C > T, predicted to impact the EEF1A1 gene expression (ß = 3.02; P-value = 3.51E-03) and the residual feed intake trait (ß = - 3.47; P-value = 0.02). This SNP was predicted to modify transcription factor sites and overlaps with several QTL for feed efficiency traits. In addition, co-expression network analyses showed that animals containing the T allele of the rs137256008 SNP may be triggering changes in the gene network. Therefore, our analyses reinforce and contribute to a better understanding of the biological mechanisms underlying gene expression control of feed efficiency traits in bovines. The cis-regulatory SNP can be used as biomarker for feed efficiency in Nelore cattle.

Ozone as an alternative decontamination process for N95 facemask and biosafety gowns.

COVID-19 pandemic created a global shortage of medical protective equipment. Here, we considered ozone (O3) a disinfectant alternative due to its potent oxidative activity against biological macromolecules. The O3 decontamination assays were done using SARS-CoV-2 obtained from patients to produce artificial contamination of N95 masks and biosecurity gowns. The quantification of SARS-CoV-2 was performed before and after exposing the samples to different ozone gas concentrations for times between 5 and 30 min. Viral loads as a function of the O3 exposure time were estimated from the data obtained by the RT-PCR technique. The genetic material of the virus was no longer detected for any tested concentrations after 15 min of O3 exposure, which means a disinfection Concentration-Time above 144 ppm min. Vibrational spectroscopies were used to follow the modifications of the polymeric fibers after the O3 treatment. The results indicate that the N95 masks could be safely reused after decontamination with treatments of 15 min at the established O3 doses for a maximum of 6 cycles.

Ex situ model of biofilm-associated wounds: providing a host-like environment for the study of Staphylococcus aureus and Pseudomonas aeruginosa biofilms.

AIM: This study aimed to assess an ex situ model of biofilm-associated wounds on porcine skin for the study of Staphylococcus aureus and Pseudomonas aeruginosa biofilms in a host-like environment, after 48 to 120 h of incubation. MATERIAL AND RESULTS: Ex situ and in vitro biofilms were comparatively analysed. Overall, CFU-counts and matrix quantification yielded significantly (P < 0·05) higher results for ex situ than in vitro biofilms. Confocal microscopy revealed greater (P < 0·05) biomass and thickness at 48-72 h and greater (P < 0·05) robustness at 72 h of growth. S. aureus ex situ biofilms produced less (P < 0·05) siderophore and proteases than in vitro biofilms, while P. aeruginosa ex situ biofilms produced more (P < 0·05) siderophores and less proteases than in vitro biofilms. CONCLUSIONS: Biofilms grown ex situ present a greater amount of bacterial cells and polymeric matrix than their in vitro counterparts, reaching maturity at 72 h of growth. Moreover the production of virulence factors differs between ex situ and in vitro biofilms. SIGNIFICANCE AND IMPACT OF THE STUDY: These findings emphasize the importance of using ex situ biofilm models, once they mimic in vivo conditions. The use of these models brings perspectives for the pursuit of therapeutic alternatives, as tests may be performed in a host-like environment.

Bessel beam optical tweezers for manipulating superparamagnetic beads.

We propose a Bessel beam optical tweezers setup that can stably trap superparamagnetic beads. The trap stiffness measured is practically independent of the radius of the Bessel beam and of the bead height (distance from the coverlip of the sample chamber), indicating that the beads can be trapped with high accuracy within a wide range of such parameters. On the other hand, the trap stiffness exhibits the expected linear increase with the laser power, despite the non-negligible absorption coefficient of the superparamagnetic beads. A geometrical optics model that considers spherical aberration and light absorption by the beads was used to predict the optical forces and trap stiffness, showing excellent agreement with the experimental data. We believe the results presented here advance the field of optical trapping manipulation of absorbing magnetic particles, and future applications will involve, for example, the design of new hybrid optomagnetic tweezers.

Olfactory response of Trichogramma pretiosum (Hymenoptera: Trichogrammatidae) to volatiles induced by transgenic maize.

Plants not only respond to herbivorous damage but adjust their defense system after egg deposition by pest insects. Thereby, parasitoids use oviposition-induced plant volatiles to locate their hosts. We investigated the olfactory behavioral responses of Trichogramma pretiosum Riley, 1879 (Hymenoptera: Trichogrammatidae) to volatile blends emitted by maize (Zea mays L.) with singular and stacked events after oviposition by Spodoptera frugiperda Smith, 1797 (Hymenoptera: Trichogrammatidae) moths. Additionally, we examined possible variations in gene expression and on oviposition-induced volatiles. We used a Y-tube olfactometer to test for the wasp responses to volatiles released by maize plants oviposited by S. frugiperda and not-oviposited plants. Using the real-time PCR technique (qRT-PCR), we analyzed the expression of lipoxygenase and three terpene synthases genes, which are enzymes involved in the synthesis of volatile compounds that attract parasitoids of S. frugiperda. Olfactometer tests showed that T. pretiosum is strongly attracted by volatiles from transgenic maize emitted by S. frugiperda oviposition (VTPRO 3, more than 75% individuals were attracted). The relative expression of genes TPS10, LOX e STC was higher in transgenic hybrids than in the conventional (isogenic line) hybrids. The GC-MS analysis revealed that some volatile compounds are released exclusively by transgenic maize. This study provides evidence that transgenic hybrids enhanced chemical cues under oviposition-induction and helped to increase T. pretiosum efficiency in S. frugiperda control. This finding shows that among the evaluated hybrids, genetically modified hybrids can improve the biological control programs, since they potentialize the egg parasitoid foraging, integrating pest management.

Transferring Occlusal Anatomy from Worn Temporary Crowns to Zirconium Oxide Crowns.

Purpose/Aim: To evaluate the accuracy of transferring the occlusal anatomy of provisional crowns to monolithic zirconium oxide crowns. Materials and Methods: From a prepared typodont-tooth (#36), ten CAD/CAM provisional polymethylethacrylate crowns were fabricated with the E4D system. Each provisional crown had its occlusion adjusted. They were scanned (E4D) and the .stl files of the crown preparations were merged with the files from the adjusted crowns (3 shape software) to produce ten polished monolithic zirconium oxide crowns. For comparison, provisional and monolithic zirconium oxide crowns were scanned (True-Definition scanner), the .stl files aligned, converted into a normalized 76x76-matrix, analyzed with ANOVA with repeated measures and Tukey's test. To generate deviation distribution tables and difference plots, .stl files (provisional crowns and monolithic zirconium oxide crowns) were merged with Geomagic software. Results: There were significant differences between provisional crowns and monolithic zirconium oxide crowns. The differences were manly in the fissure area. 86% of the calculated deviations were between + 0.06mm and - 0.04mm, 42.4% of all data points were within ± 0.022mm with a SD of 0.005mm. The main differences were in the fissures, requiring clinically none or only minimal occlusal adjustments for these zirconium oxide crowns.

On the use of Europium (Eu) for designing new metal-based anticancer drugs.

Europium oxide (Eu2O3) was used to evaluate the affinity of this rare earth element for interacting with double-stranded (ds) DNA molecules. To perform the study, we used single molecule force spectroscopy with optical tweezers and gel electrophoresis assays. Force spectroscopy experiments show that Eu2O3 presents a strong interaction with dsDNA, and the binding is independent on the ionic strength used in the surrounding environment. Among the main characteristics of the interaction, Eu2O3 tends to bind in a cooperative way, forming bound clusters of ∼ 3 molecules, and presents a high equilibrium association binding constant on the order of 105 M-1. In addition, gel electrophoresis confirm the weak electrostatic character of the interaction and explicit show that Eu2O3 does not interfere on drug intercalation into the double-helix. Such results demonstrate the potential of europium for interacting with nucleic acids and strongly suggest that this rare earth element may be considered for the design of new metal-based anticancer drugs in the future.

Silicon microparticles as handles for optical tweezers experiments.

We study the behavior of silicon microparticles in a 1064 nm Gaussian-beam optical tweezers, showing that this semiconductor can be used to perform different types of optical manipulation experiments. Depending on the focal position and the laser power used, the particles can present an oscillatory dynamics in the tweezers or can be stably 3D-trapped with a trap stiffness that allows the application of femtoNewton forces with accuracy. A new, to the best of our knowledge, interpretation based on the photoexcitation of electrons in the valence band is proposed to explain the oscillations, and the quantities associated with such dynamics (e.g., amplitude, period, etc.) were characterized as a function of relevant parameters to optical tweezers setups.

The global epidemiology of emerging Histoplasma species in recent years.

Histoplasmosis is a serious infectious disease in humans caused by Histoplasma spp. (Onygenales), whose natural reservoirs are thought to be soil enriched with bird and bat guano. The true global burden of histoplasmosis is underestimated and frequently the pulmonary manifestations are misdiagnosed as tuberculosis. Molecular data on epidemiology of Histoplasma are still scarce, even though there is increasing recognition of histoplasmosis in recent years in areas distant from the traditional endemic regions in the Americas. We used multi-locus sequence data from protein coding loci (ADP-ribosylation factor, H antigen precursor, and delta-9 fatty acid desaturase), DNA barcoding (ITS1/2+5.8s), AFLP markers and mating type analysis to determine the genetic diversity, population structure and recognise the existence of different phylogenetic species among 436 isolates of Histoplasma obtained globally. Our study describes new phylogenetic species and the molecular characteristics of Histoplasma lineages causing outbreaks with a high number of severe outcomes in Northeast Brazil between 2011 and 2015. Genetic diversity levels provide evidence for recombination, common ancestry and clustering of Brazilian isolates at different geographic scales with the emergence of LAm C, a new genotype assigned to a separate population cluster in Northeast Brazil that exhibited low diversity indicative of isolation. The global survey revealed that the high genetic variability among Brazilian isolates along with the presence of divergent cryptic species and/or genotypes may support the hypothesis of Brazil being the center of dispersion of Histoplasma in South America, possibly with the contribution of migratory hosts such as birds and bats. Outside Brazil, the predominant species depends on the region. We confirm that histoplasmosis has significantly broadened its area of occurrence, an important feature of emerging pathogens. From a practical point of view, our data point to the emergence of histoplasmosis caused by a plethora of genotypes, and will enable epidemiological analysis focused on understanding the processes that lead to histoplasmosis. Further, the description of this diversity opens avenues for comparative genomic studies, which will allow progress toward a consensus taxonomy, improve understanding of the presence of hybrids in natural populations of medically relevant fungi, test reproductive barriers and to explore the significance of this variation.

Chemical inhibition of ß-glucocerebrosidase does not affect phagocytosis and early containment of Leishmania by murine macrophages.

Gaucher disease is a lysosomal storage disease in which a genetic deficiency in ß-glucocerebrosidase leads to the accumulation of glycosphingolipids in lysosomes. Macrophages are amongst the cells most severely affected in Gaucher disease patients. One phenotype associated with Gaucher macrophages is the impaired capacity to fight bacterial infections. Here, we investigate whether inhibition of ß-glucocerebrosidase activity affects the capacity of macrophages to phagocytose and act on the early containment of human pathogens of the genus Leishmania. Towards our aim, we performed in vitro infection assays on macrophages derived from the bone marrow of C57BL/6 mice. To mimic Gaucher disease, macrophages were incubated with the ß-glucocerebrosidase inhibitor, conduritol B epoxide (CBE), prior to contact with Leishmania. This treatment guaranteed that ß-glucocerebrosidase was fully inhibited during the contact of macrophages with Leishmania, its enzymatic activity being progressively recovered along the 48 h that followed removal of the inhibitor. Infections were performed with L. amazonensis, L. infantum, or L. major, so as to explore potential species-specific responses in the context of ß-glucocerebrosidase inactivation. Parameters of infection, recorded immediately after phagocytosis, as well as 24 and 48 h later, revealed no noticeable differences in the infection parameters of CBE-treated macrophages relative to non-treated controls. We conclude that blocking ß-glucocerebrosidase activity during contact with Leishmania does not interfere with the phagocytic capacity of macrophages and the early onset of leishmanicidal responses.

Pixantrone anticancer drug as a DNA ligand: Depicting the mechanism of action at single molecule level.

In this work we use single molecule force spectroscopy performed with optical tweezers in order to characterize the complexes formed between the anticancer drug Pixantrone (PIX) and the DNA molecule, at two very different ionic strengths. Firstly, the changes of the mechanical properties of the DNA-PIX complexes were studied as a function of the drug concentration in the sample. Then, a quenched-disorder statistical model of ligand binding was used in order to determine the physicochemical (binding) parameters of the DNA-PIX interaction. In particular, we have found that the PIX molecular mechanism of action involves intercalation into the double helix, followed by a significant compaction of the DNA molecule due to partial neutralization of the phosphate backbone. Finally, this scenario of interaction was quantitatively compared to that found for the related drug Mitoxantrone (MTX), which binds to DNA with a considerably higher equilibrium binding constant and promotes a much stronger DNA compaction. The comparison performed between the two drugs can bring clues to the development of new (and more efficient) related compounds.

PDX1 -MODY and dorsal pancreatic agenesis: New phenotype of a rare disease.

Maturity-Onset Diabetes of the Young (MODY) type 4 or PDX1 -MODY is a rare form of monogenic diabetes caused by heterozygous variants in PDX1 . Pancreatic developmental anomalies related to PDX1 are reported only in neonatal diabetes cases. Here, we describe dorsal pancreatic agenesis in 2 patients with PDX1 -MODY. The proband presented with diabetes since 14 years of age and maintained regular glycemic control with low doses of basal insulin and detectable C-peptide levels after 38 years with diabetes. A diagnosis of MODY was suspected. Targeted next-generation sequencing identified a heterozygous variant in PDX1 : c.188delC/p.Pro63Argfs*60. Computed tomography revealed caudal pancreatic agenesis. Low fecal elastase indicated exocrine insufficiency. His son had impaired glucose tolerance, presented similar pancreatic agenesis, and harbored the same allelic variant. The unusual presentation in this Brazilian family enabled expansion upon a rare disease phenotype, demonstrating the possibility of detecting pancreatic malformation even in cases of PDX1 -related diabetes diagnosed after the first year of life. This finding can improve the management of MODY4 patients, leading to precocious investigation of pancreatic dysgenesis and exocrine dysfunction.

Comprehensive molecular screening strategy of OCLN in band-like calcification with simplified gyration and polymicrogyria.

Occludin (OCLN) is an important component of the tight junction complex, providing apical intercellular connections between adjacent cells in endothelial and epithelial tissue. In 2010 O'Driscoll et al reported mutations in OCLN to cause band-like calcification with simplified gyration and polymicrogyria (BLC-PMG). BLC-PMG is a rare autosomal recessive syndrome, characterized by early onset seizures, progressive microcephaly, severe developmental delay and deep cortical gray matter and basal ganglia calcification with symmetrical, predominantly fronto-parietal, polymicrogyria. Here we report 4 additional cases of BLC-PMG with novel OCLN mutations, and provide a summary of the published mutational spectrum. More generally, we describe a comprehensive molecular screening strategy taking into account the technical challenges associated with the genetic architecture of OCLN, which include the presence of a pseudo-gene and copy number variants.

Insulinoma: A retrospective study analyzing the differences between benign and malignant tumors.

BACKGROUND/OBJECTIVES: Insulinoma is a rare pancreatic tumor and, usually, a benign disease but can be a malignant one and, sometimes, a highly aggressive disease. The aim of this study was to determine differences between benign and malignant tumors. METHODS: Retrospective study of 103 patients with insulinoma treated in a tertiary center. It was analyzed demographic, clinical, laboratory, localization and histologic analysis of tumor and follow up data of subjects in order to identify differences between individuals benign and malignant disease. RESULTS: Almost all patients (87%) had a benign tumor and survival rates of 100% following pancreatic tumor surgery. Those with malignant tumors (13%) have a poor prognosis, 77% insulinoma-related deaths over a period of 1-300 months after the diagnosis with a survival rate of 24% in five years. The following factors are associated with an increased risk of malignant disease: duration of symptoms < 24 months, fasting time for the occurrence of hypoglycemia < 8 h, blood plasma insulin concentration ≥ 28 µU/mL and C-peptide ≥ 4.0 ng/mL at the glycemic nadir and tumor size ≥ 2.5 cm. CONCLUSIONS: Our data help to base the literature about these tumors, reinforcing that although insulinoma is usually a single benign and surgically treated neoplasia, the malignant one is difficult to treat. We highlight the data that help predict a malignancy behavior of tumor and suggest a long follow up after diagnosis in these cases.

A cooperative transition from the semi-flexible to the flexible regime of polymer elasticity: Mitoxantrone-induced DNA condensation.

We report a high cooperative transition from the semi-flexible to the flexible regime of polymer elasticity during the interaction of the DNA molecule with the chemotherapeutic drug Mitoxantrone (MTX). By using single molecule force spectroscopy, we show that the force-extension curves of the DNA-MTX complexes deviate from the typical worm-like chain behavior as the MTX concentration in the sample increases, becoming straight lines for sufficiently high drug concentrations. The behavior of the radius of gyration of the complexes as a function of the bound MTX concentration was used to quantitatively investigate the cooperativity of the condensation process. The present methodology can be promptly applied to other ligands that condense the DNA molecule upon binding, opening new possibilities in the investigation of this type of process and, more generally, in the investigation of phase transitions in polymer physics.