scholarly journals Electrochemical DNA Biosensor That Detects Early Celiac Disease Autoantibodies

Sensors ◽  
2021 ◽  
Vol 21 (8) ◽  
pp. 2671
Author(s):  
Anna B. N. Nguyen ◽  
Marcos Maldonado ◽  
Dylan Poch ◽  
Tyler Sodia ◽  
Andrew Smith ◽  
...  
Keyword(s):  
Celiac Disease ◽  
Dna Biosensor ◽  
Diagnostic Tools ◽  
Dependent Manner ◽  
Apparent Dissociation Constant ◽  
Specific Patient ◽  
Invasive Method ◽  
Target Binding ◽  
Current Reduction ◽  
Invasive Diagnostics

Although it is estimated that more than one million Americans have celiac disease (CD), it remains challenging to diagnose. CD, an autoimmune and inflammatory response following the ingestion of gluten-containing foods, has symptoms overlapping with other diseases and requires invasive diagnostics. The gold standard for CD diagnosis involves serologic blood tests followed by invasive confirmatory biopsies. Here, we propose a less invasive method using an electrochemical DNA (E-DNA) biosensor for CD-specific autoantibodies (AABs) circulating in blood. In our approach, CD-specific AABs bind a synthetic neoepitope, causing a conformational change in the biosensor, as well as a change in the environment of an attached redox reporter, producing a measurable current reduction. We assessed the biosensor’s ability to detect CD-specific patient-derived AABs in physiological buffer as well as buffer supplemented with bovine serum. Our biosensor was able to detect AABs in a dose-dependent manner; increased signal change correlated with increased AAB concentration with an apparent dissociation constant of 0.09 ± 0.03 units/mL of AABs. Furthermore, we found our biosensor to be target-specific, with minimal off-target binding of multiple unrelated biomarkers. Future efforts aimed at increasing sensitivity in complex media may build upon the biosensor design presented here to further improve CD AAB detection and CD diagnostic tools.

scholarly journals Tools Used to Measure the Physical State of Women with Celiac Disease: A Review with a Systematic Approach

2020 ◽  
Vol 17 (2) ◽  
pp. 539
Author(s):  
Alejandro Martínez-Rodríguez ◽  
Daniela Alejandra Loaiza-Martínez ◽  
Javier Sánchez-Sánchez ◽  
Pablo J. Marcos-Pardo ◽  
Soledad Prats ◽  
...  
Keyword(s):  
Celiac Disease ◽  
Selection Process ◽  
Immunoglobulin A ◽  
Clinical History ◽  
Diagnostic Tools ◽  
Intestinal Biopsy ◽  
Specific Protocol ◽  
Small Intestinal Biopsy ◽  
Immunological Disorder ◽  
Meta Analyses

Celiac disease (CD) is an immunological disorder that mainly affects the small intestine, generating an inflammatory process in response to the presence of gluten (a protein). Autoimmune diseases are part of a group of diseases that are difficult to diagnose without a specific protocol or consensus to detect them due to the number of symptoms and diseases with which it has a relationship. Therefore, the aim of this review was to analyze the diagnostic tools of CD used in middle-aged women, to compare the use and effectiveness of the different tools, and to propose a strategy for the use of the tools based on the results found in the literature. The present research followed the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guideline. The search was conducted in the following databases: Scielo, PubMed, Web of Science, and Worldwide Science org. In the initial literature search, 2004 titles and relevant abstracts were found. Among them, 687 were duplicates, leaving 1130 articles. Based on the inclusion criteria, only 41 articles passed the selection process; 4 main types of analyses appear in the studies: blood tests, questionnaires, clinical history, and biopsy. It can be said that none of the analyses have a 100% reliability since most of them can present false negatives; therefore, the best way to diagnose celiac disease up to now is through a combination of different tests (Immunoglobulin A and small intestinal biopsy).

scholarly journals Charybdotoxin selectively blocks small Ca-activated K channels in Aplysia neurons.

1987 ◽  
Vol 90 (1) ◽  
pp. 27-47 ◽  
Author(s):  
A Hermann ◽  
C Erxleben
Keyword(s):  
Single Channel ◽  
Delayed Rectifier ◽  
Pacemaker Activity ◽  
Dependent Manner ◽  
Open Probability ◽  
Apparent Dissociation Constant ◽  
External Application ◽  
K Channels ◽  
Voltage Dependent ◽  
K Current

The action of charybdotoxin (ChTX), a peptide component isolated from the venom of the scorpion Leiurus quinquestriatus, was investigated on membrane currents of identified neurons from the marine mollusk, Aplysia californica. Macroscopic current recordings showed that the external application of ChTX blocks the Ca-activated K current in a dose- and voltage-dependent manner. The apparent dissociation constant is 30 nM at V = -30 mV and increases e-fold for a +50- to +70-mV change in membrane potential, which indicates that the toxin molecule is sensitive to approximately 35% of the transmembrane electric field. The toxin is bound to the receptor with a 1:1 stoichiometry and its effect is reversible after washout. The toxin also suppresses the membrane leakage conductance and a resting K conductance activated by internal Ca ions. The toxin has no significant effect on the inward Na or Ca currents, the transient K current, or the delayed rectifier K current. Records from Ca-activated K channels revealed a single channel conductance of 35 +/- 5 pS at V = 0 mV in asymmetrical K solution. The channel open probability increased with the internal Ca concentration and with membrane voltage. The K channels were blocked by submillimolar concentrations of tetraethylammonium ions and by nanomolar concentrations of ChTX, but were not blocked by 4-aminopyridine if applied externally on outside-out patches. From the effects of ChTX on K current and on bursting pacemaker activity, it is concluded that the termination of bursts is in part controlled by a Ca-activated K conductance.

scholarly journals Ca2+-dependent and phospholipid-independent binding of annexin 2 and annexin 5

2002 ◽  
Vol 367 (3) ◽  
pp. 895-900 ◽  
Author(s):  
Nicole D. BROOKS ◽  
Jean E. GRUNDY ◽  
Nadine LAVIGNE ◽  
Mélanie C. DERRY ◽  
Christina M. RESTALL ◽  
...  
Keyword(s):  
Binding Sites ◽  
Dependent Manner ◽  
Homologous Proteins ◽  
Anionic Phospholipid ◽  
Apparent Dissociation Constant ◽  
Excess Amount ◽  
Annexin 2 ◽  
Covalently Linked ◽  
Reporter Group ◽  
Dissociation Constant Kd

Annexins are a family of homologous proteins that associate with anionic phospholipid (aPL) in the presence of Ca2+. Evidence that the function of one annexin type may be regulated by another was recently reported in studies investigating cytomegalovirus—aPL interactions, where the fusogenic function of annexin 2 (A2) was attenuated by annexin 5 (A5). This observation suggested that A2 may bind directly to A5. In the present study, we demonstrated this interaction. The A2—A5 complex was first detected utilizing (covalently linked) fluorescein-labelled A5 (F-A5) as a reporter group. The interaction required concentrations of Ca2+ in the millimolar range, had an apparent dissociation constant [Kd(app)] of 1nM at 2mM Ca2+ and was independent of aPL. A2 bound comparably with F-A5 pre-equilibrated with an amount of aPL that could bind just the F-A5 or to an excess amount of aPL providing sufficient binding sites for all of F-A5 and A2. A2—A5 complex formation was corroborated in an experiment, where [125I]A2 associated in a Ca2+-dependent manner with A5 coated on to polystyrene. Surface plasmon resonance was used as a third independent method to demonstrate the binding of A2 and A5 and, furthermore, supported the conclusion that the monomeric and tetrameric forms of A2 bind equivalently to A5. Together these results demonstrate an A2—A5 interaction and provide an explanation as to how A5 inhibits the previously reported A2-dependent enhancement of virus—aPL fusion.

scholarly journals Identification of Aptamers that Specifically Bind to A1 Antigen by Performing Cell-on Human Erythrocytes

2020 ◽  
Vol 9 ◽  
pp. 1657
Author(s):  
Seyed Mohammad Hasan Hosseini ◽  
Mohammad Reza Bassami ◽  
Alireza Haghparast ◽  
Mojtaba Sankian ◽  
Gholamreza Hashemi Tabar
Keyword(s):  
Blood Group ◽  
Dna Aptamer ◽  
Diagnostic Tools ◽  
Blood Group Antigens ◽  
Apparent Dissociation Constant ◽  
Fluorescent Intensity ◽  
Target Molecules ◽  
Polymerase Chain ◽  
Dissociation Constant Kd

Background: The apply of aptamers as a new generation’s way to probe diagnostic for the detection of target molecules has gained ground. Aptamers can be used as alternatives to diagnostic antibodies for detection of blood groups due to their unique features. This study was aimed to produce DNA diagnostic aptamer detecting the antigen of A1 blood group using the Cell-Selex method. Materials and Methods: DNA aptamer was isolated against A1 RBC antigen after ten stages of Cell-Selex and amplification by an asymmetric polymerase chain reaction. The progress of the stages of selection was evaluated using flow cytometry analysis, which the DNA aptamer isolated from the tenth cycle with an affinity of 70% fluorescent intensity, was selected from four positive colonies followed by determination of the sequences and secondary structures. Results: The aptameric sequence obtained from C4 cloning was calculated with the highest binding affinity to A1 antigen having an apparent dissociation constant (Kd value) of at least 29.5 ± 4.3 Pmol, which was introduced as the selected aptamer-based on ΔG obtained from a colony of C4 equal to –13.13. Conclusion: The aptamer obtained from using Cell-Selex method could be used as an example for the development of diagnostic tools such as biosensors for detecting A1 blood group antigens. [GMJ.2020;9:e1657] 

scholarly journals The Evolving Landscape of Biomarkers in Celiac Disease: Leading the Way to Clinical Development

2021 ◽  
Vol 12 ◽  
Author(s):  
Glennda Smithson ◽  
Jenifer Siegelman ◽  
Toshihiko Oki ◽  
Joseph R. Maxwell ◽  
Daniel A. Leffler
Keyword(s):  
Celiac Disease ◽  
Peripheral Blood Cell ◽  
Clinical Development ◽  
Intestinal Damage ◽  
Gluten Free ◽  
Histological Changes ◽  
Specific Patient ◽  
Cell Responses ◽  
Immune Mediated ◽  
Duodenal Biopsies

Celiac disease is a common immune-mediated disease characterized by abnormal T-cell responses to gluten. For many patients, symptoms and intestinal damage can be controlled by a gluten-free diet, but, for some, this approach is not enough, and celiac disease progresses, with serious medical consequences. Multiple therapies are now under development, increasing the need for biomarkers that allow identification of specific patient populations and monitoring of therapeutic activity and durability. The advantage of identifying biomarkers in celiac disease is that the underlying pathways driving disease are well characterized and the histological, cellular, and serological changes with gluten response have been defined in gluten challenge studies. However, there is room for improvement. Biomarkers that measure histological changes require duodenal biopsies and are invasive. Less invasive peripheral blood cell and cytokine biomarkers are transient and dependent upon gluten challenge. Here, we discuss established biomarkers and new approaches for biomarkers that may overcome current limitations.

scholarly journals Batrachotoxin-modified sodium channels from squid optic nerve in planar bilayers. Ion conduction and gating properties.

1989 ◽  
Vol 93 (1) ◽  
pp. 23-41 ◽  
Author(s):  
M I Behrens ◽  
A Oberhauser ◽  
F Bezanilla ◽  
R Latorre
Keyword(s):  
Optic Nerve ◽  
Dissociation Constant ◽  
Sodium Channels ◽  
Single Channel ◽  
Dependent Manner ◽  
Channel Conductance ◽  
Apparent Dissociation Constant ◽  
Planar Bilayers ◽  
Voltage Range ◽  
Voltage Dependent

Squid optic nerve sodium channels were characterized in planar bilayers in the presence of batrachotoxin (BTX). The channel exhibits a conductance of 20 pS in symmetrical 200 mM NaCl and behaves as a sodium electrode. The single-channel conductance saturates with increasing the concentration of sodium and the channel conductance vs. sodium concentration relation is well described by a simple rectangular hyperbola. The apparent dissociation constant of the channel for sodium is 11 mM and the maximal conductance is 23 pS. The selectivity determined from reversal potentials obtained in mixed ionic conditions is Na+ approximately Li+ greater than K+ greater than Rb+ greater than Cs+. Calcium blocks the channel in a voltage-dependent manner. Analysis of single-channel membranes showed that the probability of being open (Po) vs. voltage relation is sigmoidal with a value of 0.5 between -90 and -100 mV. The fitting of Po requires at least two closed and one open state. The apparent gating charge required to move through the whole transmembrane voltage during the closed-open transition is four to five electronic charges per channel. Distribution of open and closed times are well described by single exponentials in most of the voltage range tested and mean open and mean closed times are voltage dependent. The number of charges associated with channel closing is 1.6 electronic charges per channel. Tetrodotoxin blocked the BTX-modified channel being the blockade favored by negative voltages. The apparent dissociation constant at zero potential is 16 nM. We concluded that sodium channels from the squid optic nerve are similar to other BTX-modified channels reconstituted in bilayers and to the BTX-modified sodium channel detected in the squid giant axon.

Albumin endocytosis is regulated by heterotrimeric GTP-binding protein G alpha i-3 in opossum kidney cells

1996 ◽  
Vol 271 (2) ◽  
pp. F356-F364 ◽  
Author(s):  
N. J. Brunskill ◽  
N. Cockcroft ◽  
S. Nahorski ◽  
J. Walls
Keyword(s):  
G Protein ◽  
Pertussis Toxin ◽  
Protein G ◽  
Dependent Manner ◽  
Protein Subunit ◽  
Apparent Dissociation Constant ◽  
Opossum Kidney ◽  
Ok Cells ◽  
Proximal Tubular ◽  
Albumin Endocytosis

Proteinuria is an adverse feature in patients with renal disease, possibly due to toxicity of albumin to proximal tubular cells. Albumin is reabsorbed from tubular fluid by receptor-mediated endocytosis. The mechanism of regulation of the endocytosis is unknown. The large quantities of G proteins in proximal tubular cell apical membranes suggests that they may have a regulatory role in endocytosis. 125I-labeled albumin uptake was measured in opossum kidney (OK) cells. This is a saturable process with high-affinity [apparent dissociation constant (Kd) = 24.3 mg/l] and low-affinity (Kd = 15.9 g/l) components. The endocytic uptake of gold-albumin into OK cells was confirmed by electron microscopy. 125I-albumin endocytosis in OK cells was inhibited by pertussis toxin, but cholera toxin had no effect. Pertussis toxin also inhibited uptake of [3H]inulin. OK cells were stably transfected with a cDNA for the G protein subunit G alpha i-3 and transfectants were screened by immunoblotting. Several G alpha i-3-overexpressing clones were detected. OK cells overexpressing G alpha i-3 demonstrate increased 125I-albumin uptake, which is abolished by pertussis toxin, in both a concentration- and time-dependent manner. These results suggest that albumin endocytosis in OK cells is regulated by the G protein G alpha i-3.

P2X receptors in mouse Leydig cells

2006 ◽  
Vol 290 (4) ◽  
pp. C1009-C1017 ◽  
Author(s):  
Luiz Artur Poletto Chaves ◽  
Endrigo Piva Pontelli ◽  
Wamberto Antonio Varanda
Keyword(s):  
Leydig Cells ◽  
Pyridoxal Phosphate ◽  
Hill Coefficient ◽  
Disulfonic Acid ◽  
Inward Rectification ◽  
Dependent Manner ◽  
Patch Clamp Technique ◽  
Concentration Dependent Manner ◽  
Apparent Dissociation Constant ◽  
Whole Cell

ATP-activated currents were studied in Leydig cells of mice with the patch-clamp technique. Whole cell currents were rapidly activating and slowly desensitizing (55% decrement from the peak value on exposure to 100 μM ATP for 60 s), requiring 3 min of washout to recover 100% of the response. The concentration-response relationships for ATP, adenosine 5′- O-(3-thiotriphosphate) (ATPγS), and 2-methylthio-ATP (2-MeS-ATP) were described by the Hill equation with a concentration evoking 50% of maximal ATP response ( Kd) of 44, 110, and 637 μM, respectively, and a Hill coefficient of 2. The order of efficacy of agonists was ATP ≥ ATPγS > 2-MeS-ATP > 2′,3′- O-(4-benzoylbenzoyl)-ATP (BzATP). αβ-Methylene-ATP (αβ-MeATP), GTP, UTP, cAMP, and adenosine were ineffective. Suramin and pyridoxal phosphate-6-azophenyl-2′,4′-disulfonic acid (PPADS) blocked the responses in a concentration-dependent manner. The ATP-activated currents were dependent on extracellular pH, being maximal at pH 6.5 and decreasing with both acidification and alkalinization (apparent dissociation constant (p Ka) of 5.9 and 7.4, respectively). The whole cell current-voltage relationship showed inward rectification and reversed near 0 mV. Experiments performed in bi-ionic conditions for measurement of reversal potentials showed that this channel is highly permeable to calcium [permeability ( P)Ca/ PNa = 5.32], but not to chloride ( PCl/ PNa = 0.03) or N-methyl-d-glucamine (NMDG) ( PNMDG/ PNa = 0.09). Unitary currents recorded in outside-out patches had a chord conductance of 27 pS (between −90 and −50 mV) and were inward rectifying. The average current passing through the excised patch decreased with time [time constant (τ) = 13 s], resembling desensitization of the macroscopic current. These findings indicate that the ATP receptor present in Leydig cells shows properties most similar to those of cloned homomeric P2X2.

scholarly journals Calcium current activated upon hyperpolarization of Paramecium tetraurelia.

1992 ◽  
Vol 100 (2) ◽  
pp. 233-251 ◽  
Author(s):  
R R Preston ◽  
Y Saimi ◽  
C Kung
Keyword(s):  
Dissociation Constant ◽  
Calcium Current ◽  
Divalent Cations ◽  
Dependent Manner ◽  
Paramecium Tetraurelia ◽  
Apparent Dissociation Constant ◽  
Voltage Dependent ◽  
Effective Valence ◽  
The One ◽  
K Currents

Hyperpolarization of Paramecium tetraurelia under conditions where K+ currents are suppressed elicits an inward current that activates rapidly toward a peak at 25-80 ms and decays thereafter. This peak current (Ihyp) is not affected by removing Cl ions from the microelectrodes used to clamp membrane potential, or by changing extracellular Cl- concentration, but is lost upon removing extracellular Ca2+. Ihyp is also lost upon replacing extracellular Ca2+ with equimolar concentrations of Ba2+, Co2+, Mg2+, Mn2+, or Sr2+, suggesting that the permeability mechanism that mediates Ihyp is highly selective for Ca2+. Divalent cations also inhibit Ihyp when introduced extracellularly, in a concentration- and voltage-dependent manner. Ba2+ inhibits Ihyp with an apparent dissociation constant of 81 microM at -110 mV, and with an effective valence of 0.42. Ihyp is also inhibited reversibly by amiloride, with a dissociation constant of 0.4 mM. Ihyp is not affected significantly by changes in extracellular Na+, K+, or H+ concentration, or by EGTA injection. Also, it is unaffected by manipulations or mutations that suppress the depolarization-activated Ca2+ current or the various Ca(2+)-dependent currents of Paramecium. We suggest that Ihyp is mediated by a novel, hyperpolarization-activated calcium conductance that is distinct from the one activated by depolarization.

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