Purpose: To analyze the morphological outcomes of the posterior corneal opacity or "semilunar sign" in noninfectious anterior scleritis using multimodal imaging. Methods: This was a prospective ...observational case series. Patients with anterior scleritis from January 2018 to January 2019 were included. Clinical and demographic data were collected. Posterior cornea was visualized using the digital slit lamp photography (Elite, mega digital vision), spectral domain optical coherence tomography (MS39), and specular count analyzer (EM-3000). "Semilunar sign" was defined by the (1) presence of posterior corneal opacity, (2) concave semilunar pattern, (3) absence of blood vessels, and (4) normal anterior cornea. Incidence, clinical characteristics and significance, correlation with Mantoux sensitivity, and role of multimodal valuation were assessed. Results: Overall 76 eyes of 72 patients were recruited with anterior scleritis. Fifteen eyes of 11 patients (15.3%) presented with semilunar sign. The scleritis was both nonnecrotizing (n = 8) and necrotizing (n = 7). The semilunar configuration appeared as isolated (n = 9) and continuous lesion (n = 6). The extent was directly related to the scleral disease extent (P = 0.002). The mean thickness measured 212.5 ± 129.3 μm. The mean central endothelial cell density (ECD) was 2540.8 ± 351.7 cells/mm2, which was significantly higher than the involved peripheral cornea (P = 0.05). The mean surface area of the semilunar sign was 7.7 ± 5.2 mm2. There was no significant correlation between the opacity thickness and the best-corrected visual acuity (P = 0.895, r = −0.39), ECD (P = 0.52, r = −0.188), and Mantoux (P = 0.696, r =− 0.142). Conclusion: Corneal semilunar sign of scleritis affected the peripheral cornea and caused no functional abnormality in early presentation. Multimodal analysis can aid in clinical assessment and severity.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Aptamers are RNA/DNA oligonucleotide molecules that specifically bind to a targeted complementary molecule. As potential recognition elements with promising diagnostic and therapeutic applications, ...aptamers, such as monoclonal antibodies, could provide many treatment and diagnostic options for blood diseases. Aptamers present several superior features over antibodies, including a simple in vitro selection and production, ease of modification and conjugation, high stability, and low immunogenicity. Emerging as promising alternatives to antibodies, aptamers could overcome the present limitations of monoclonal antibody therapy to provide novel diagnostic, therapeutic, and preventive treatments for blood diseases. Researchers in several biomedical areas, such as biomarker detection, diagnosis, imaging, and targeted therapy, have widely investigated aptamers, and several aptamers have been developed over the past two decades. One of these is the pegaptanib sodium injection, an aptamer-based therapeutic that functions as an anti-angiogenic medicine, and it is the first aptamer approved by the U.S. Food and Drug Administration (FDA) for therapeutic use. Several other aptamers are now in clinical trials. In this review, we highlight the current state of aptamers in the clinical trial program and introduce some promising aptamers currently in pre-clinical development for blood diseases.
The work describes a simple cotton swab-based colorimetric immunoassay as a rapid screening tool for pathogenic bacteria on poultry processing plants. This immunosensing platform can be used for the ...detection of pathogens present on surfaces such as glass, stainless steel and chicken meat. Unlike the reported assays, here, cotton swab plays dual function: as a sample collector from the solid surfaces and as detection platform. The immunoassay was tested for the detection of 4 different bacteria;
Salmonella typhimurium, Salmonella enteritidis, Staphylococcus aureus
and
Campylobacter jejuni
. The immunoassays were fabricated by immobilizing specific antibody for each bacterium on a cotton swab that is used to recover the cells from contaminated surfaces. Then, a sandwich immunoassay was developed by immersing the cotton swab in different colored nanobead-conjugated antibody solutions which corresponds to different bacteria. The immunoassays response was detected colorimetrically by following the change in the color intensity produced by the nanobeads due to the specific binding on the cotton swab. This simple colorimetric assay is very sensitive with a detection limit of 10 cfu.mL
−1
. Furthermore, no significant cross reactivity of the immunoassays with non specific bacteria was observed indicating good selectivity of the immunoassays. This simple, disposable and easy-to- use colorimetric platform shows great promise as rapid qualitative and semi quantitative detection tool for microorganisms on food processing plants and other surfaces.
Graphical abstract
Schematic of the sandwich colorimetric immunosensor for the detection of pathogenic bacteria on poultry processing plants using cotton swabs and nanobeads.
▸ A stable, label-free, bacteriophage-based detection of E. coli using ultra sensitive long-period gratings is demonstrated. ▸ E. coli binding by covalently immobilized bacteriophage T4 is ...investigated using the spectral interrogation mechanism. ▸ No moving part or metallization is required in our sensor, making it extremely accurate, very compact and cost effective. ▸ The detection mechanism is capable of reliable detection of E. coli concentrations as low as 103cfu/ml with an experimental accuracy > 99%. ▸ We present the SEM images to confirm a reliable binding of E. coli bacteria to the optical fiber surface.
In this paper we report a stable, label-free, bacteriophage-based detection of Escherichia coli (E. coli) using ultra sensitive long-period fiber gratings (LPFGs). Bacteriophage T4 was covalently immobilized on optical fiber surface and the E. coli binding was investigated using the highly accurate spectral interrogation mechanism. In contrast to the widely used surface plasmon resonance (SPR) based sensors, no moving part or metal deposition is required in our sensor, making the present sensor extremely accurate, very compact and cost effective. We demonstrated that our detection mechanism is capable of reliable detection of E. coli concentrations as low as 103cfu/ml with an experimental accuracy greater than 99%.
Cylindrospermopsin (CYN) is one of the most concerning cyanotoxins due to its potential toxicity and spreading to various environments including drinking water. CYN has potential interferences with ...human and animal metabolic pathways, which influence the functions of organs including liver, kidneys, lungs, etc. CYN is involved in the inhibition of protein synthesis and detachment of ribosomes from the endoplasmic reticulum membrane. It also interacts with soluble proteins, which are associated with protein translations. It is believed that cytochrome 450 is responsible for the rapid toxicity of CYN. Researchers are urged to develop a high-throughput screening method for the detection of CYN in water. Construction of low cost, rapid, and sensitive analytical methods for the detection of CYN is challenging. Here, we used graphene oxide (GO) as the fluorescence sensing platform for probing the high affinity of the short aptamer derived from the wild-type long aptamer-CYN sensing. The biosensor construction involved two steps: first, quenching the fluorescence of fluorescent-labelled truncated aptamer using GO as a quencher and, second, fluorescence recovery in the presence of CYN by competitive binding between the target and GO. One of the truncate aptamers has a 12-fold higher affinity and enhances sensitivity compared to the long aptamer sequence. The limit of detection of the high affinity truncated aptamer is 17 pM which is 6-fold lower than the long aptamer (100 pM). The sensor specifically detects CYN in the presence of other potential interfering toxins. The performance of the sensor was validated using CYN spiked tap water with very good recovery percentage. A rapid and highly sensitive detection of CYN from water resources has been achieved using this method.
Aptamers are short single-stranded oligonucleotides (either DNA or RNA) that can fold into well-defined three-dimensional (3D) spatial structures which enable them to capture their specific target by ...complementary shape interactions. Aptamers are selected from large random libraries through the SELEX process and only a small fraction of the sequence is involved in direct docking with the target. In this paper, we describe the possible truncation variants of zearalenone (ZEA) aptamer which might be an effective binding region for the target. The originally selected zearalenone (ZEA) aptamer was 80-mer in length and shown to bind the target with a high affinity (
K
d
= 41 ± 5 nM). Herein, computational docking simulation was performed with 15 truncated variants to determine the predicted binding energy and responsible binding site of the aptamer-analyte complex. The results revealed that 5 truncated variants had binding energy lower than − 7.0 kcal/mol. Circular dichroism analysis was performed on the shortlisted aptamer and the conformational change of aptamers was observed with the presence of an analyte. Aptamer Z3IN (29-mer) was chosen as the most enhanced affinity for its target with a dissociation constant of 11.77 ± 1.44 nM. The aptamer was further applied in the electrochemical aptasensor of ZEA based on an indirect competitive format. The results demonstrated that the truncated aptamer leads to an enhancement of the sensitivity of the biosensor.
Graphical abstract
An electrochemical aptasensor is described for determination of the phytohormone of zearalenone (ZEA). The gold electrode was modified with ZEA via covalent attachment using cysteamine-hydrochloride ...and 1,4-phenylene diisocyanate linker. A truncated ZEA aptamer with a dissociation constant of 13.4 ± 2.1 nM was used in an aptasensor. The electrochemical property was investigated using square wave voltammetry for monitoring the change in the electron transfer using the ferro/ferricyanide system as redox probe. Under optimal experimental conditions, the response was best measured at a potential of 0.20 V (vs. Ag/AgCl). The signals depended on the competitive mechanism between the immobilised ZEA and free ZEA for the aptamer binding site. The aptasensor works in the range 0.01 to 1000 ng·mL
−1
ZEA concentration, with a detection limit of 0.017 ng·mL
−1
. High degree of cross-reactivity with the other analogues of ZEA was observed, whereas none towards other mycotoxins. The aptasensor was further applied for the determination of ZEA in the extract of maize grain and showed good recovery percentages between 87 and 110%.
Graphical abstract
Schematic representation of the electrochemical determination of zearalenone based on indirect competitive assay. Step
a
Immobilisation of ZEA on the surface of gold electrode via covalent attachment,
b
competition for the ZEA aptamer binding site between immobilised and free ZEA, and
c
current signal of the binding event based on SWV technique.
Snakebite is a neglected tropical disease that causes considerable death and disability in the tropical world. Although snakebite can cause a variety of pathologies in victims, haemotoxic effects are ...particularly common and are typically characterised by haemorrhage and/or venom-induced consumption coagulopathy. Antivenoms are the mainstay therapy for treating the toxic effects of snakebite, but despite saving thousands of lives annually, these therapies are associated with limited cross-snake species efficacy due to venom variation, which ultimately restricts their therapeutic utility to particular geographical regions. In this study, we sought to explore the potential of ssDNA aptamers as toxin-specific inhibitory alternatives to antibodies. As a proof of principle model, we selected snake venom serine protease toxins, which are responsible for contributing to venom-induced coagulopathy following snakebite envenoming, as our target. Using SELEX technology, we selected ssDNA aptamers against recombinantly expressed versions of the fibrinogenolytic SVSPs ancrod from the venom of C. rhodostoma and batroxobin from B. atrox. From the resulting pool of specific ssDNA aptamers directed against each target, we identified candidates that exhibited low nanomolar binding affinities to their targets. Downstream aptamer-linked immobilised sorbent assay, fibrinogenolysis, and coagulation profiling experiments demonstrated that the candidate aptamers were able to recognise native and recombinant SVSP toxins and inhibit the toxin- and venom-induced prolongation of plasma clotting times and the consumption of fibrinogen, with inhibitory potencies highly comparable to commercial polyvalent antivenoms. Our findings demonstrate that rationally selected toxin-specific aptamers can exhibit broad in vitro cross-reactivity against toxin isoforms found in different snake venoms and are capable of inhibiting toxins in pathologically relevant in vitro and ex vivo models of venom activity. These data highlight the potential utility of ssDNA aptamers as novel toxin-inhibiting therapeutics of value for tackling snakebite envenoming.
MicroRNAs (miRNAs) are short non-coding RNAs that are found in various cellular compartments and play an important role in regulating gene expression. Extracellular miRNAs, such as those found within ...extracellular vesicles such as exosomes are involved in cell-to-cell communication. The intercellular transfer of miRNAs has been implicated in various diseases’ pathogenesis including cancer and has been studied extensively as potential cancer biomarkers. However, the extraction of miRNA from exosomes is still a challenging task. The current nucleic acid extraction assays are expensive and labor-intensive. In this study, we demonstrated a microfluidic device for aptamer-based magnetic separation of the exosomes and subsequent detection of the miRNA using a fluorescence switching assay, which was enabled by carbon nanomaterials coated on magnetic beads. In the OFF state, the fluorophore-labelled cDNA is quenched using carbon nanomaterials. However, when the target miRNA210 is introduced, the cDNA detaches from the bead’s surface, which leads to an increase in the fluorescence intensity (ON state). This increment was found to be proportional to miRNA concentration within the dynamic range of 0–100 nM with a detection limit of 5 pM. The assay was validated with spiked miRNA using the standard RT-PCR method. No notable cross-reactivity with other closely related miRNAs was observed. The developed method can be utilized for the minimally invasive detection of cancer biomarkers.
The novel corona (SARS-CoV-2) virus causes a global pandemic, which motivates researchers to develop reliable and effective methods for screening and detection of SARS-CoV-2. Though there are several ...methods available for the diagnosis of SARS-CoV-2 such as RT-PCR and ELSIA, nevertheless, these methods are time-consuming and may not apply at the point of care. In this study, we have developed a specific, sensitive, quantitative and fast detection method for SARS-CoV-2 by fluorescence resonance energy transfer (FRET) assay. The total extracellular protease proteolytic activity from the virus has been used as the biomarker. The specific peptide sequences from the library of 115 dipeptides were identified via changes in the fluorescence signal. The fluorogenic dipeptide substrates have the fluorophore and a quencher at the N- and the C- terminals, respectively. When the protease hydrolyzes the peptide bond between the two specific amino acids, it leads to a significant increase in the fluorescence signals. The specific fluorogenic peptide (H-d) produces a high fluorescence signal. A calibration plot was obtained from the changes in the fluorescence intensity against the different concentrations of the viral protease. The lowest limit of detection of this method was 9.7 ± 3 pfu/mL. The cross-reactivity of the SARS-CoV-2-specific peptide was tested against the MERS-CoV which does not affect the fluorescence signal. A significant change in the fluorescence signal with patient samples indicates that this FRET-based assay might be applied for the diagnosis of SARS-CoV-2 patients.
Graphical abstract
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