scholarly journals Precision localization of cellular proteins with fluorescent Fab-based probes

2021 ◽  
Author(s):  
Federica Liccardo ◽  
Matteo Lo Monte ◽  
Brunella Corrado ◽  
Martina Veneruso ◽  
Simona Celentano ◽  
...  
Keyword(s):  
Unmet Need ◽  
Super Resolution ◽  
Cellular Protein ◽  
Antigen Binding ◽  
Fab Fragment ◽  
Technical Limitation ◽  
Efficient Detection ◽  
Wide Range ◽  
Linkage Error ◽  
Fragment Antigen Binding

Currently, a major technical limitation of microscopy based image analysis is the linkage error – which describes the distance between e.g. the target epitope of cellular protein to the fluorescence emitter, which position is finally detected in a microscope. With continuously improving resolution of today′s (super–resolution) microscopes, the linkage errors can severely hamper the correct interpretation of images and is usually introduced in experiments by the use of standard intracellular staining reagents such as fluorescently labelled antibodies. The linkage error of standard labelled antibodies is caused by the size of the antibody and the random distribution of fluorescent emitters on the antibody surface. Together, these two factors account for a fluorescence displacement of ~40nm when staining proteins by indirect immunofluorescence; and ~20nm when staining with fluorescently coupled primary antibodies. In this study, we describe a class of staining reagents that effectively reduce the linkage error by more than five–fold when compared to conventional staining techniques. These reagents, called Fluo–N–Fabs, consist of an antigen binding fragment of a full-length antibody (Fab / fragment antigen binding) that is selectively conjugated at the N-terminal amino group with fluorescent organic molecules, thereby reducing the distance between the fluorescent emitter and the protein target of the analysis. Fluo–N–Fabs also exhibit the capability to penetrate tissues and highly crowded cell compartments, thus allowing for the efficient detection of cellular epitopes of interest in a wide range of fixed samples. We believe this class of reagents realize an unmet need in cell biological super resolution imaging studies where the precise localization of the target of interest is crucial for the understanding of complex biological phenomena.

scholarly journals Biofunctionalized Two-dimensional MoS2 Receptors for Rapid Response Modular Electronic SARS-CoV-2 and Influenza A Antigen Sensors

2020 ◽  
Author(s):  
C. Muratore ◽  
M.K. Muratore ◽  
D.R. Austin ◽  
P. Look ◽  
A.K. Benton ◽  
...  
Keyword(s):  
Influenza A ◽  
Antibody Fragment ◽  
Antibody Fragments ◽  
Thin Layers ◽  
Receptor Protein ◽  
Antigen Binding ◽  
Two Dimensional ◽  
Fab Fragment ◽  
Fragment Antigen Binding ◽  
Transducer Surface

AbstractMultiplex electronic antigen sensors for detection of SARS-Cov-2 spike glycoproteins or hemagglutinin from Influenza A in liquid samples with characteristics resembling extracted saliva were fabricated using scalable processes with potential for economical mass-production. The sensors utilize the sensitivity and surface chemistry of a two-dimensional MoS2 transducer for attachment of antibody fragments in a conformation favorable for antigen binding. Ultra-thin layers (3 nm) of amorphous MoS2 were directly sputtered over the entire sensor chip at room temperature and laser annealed to create an array of semiconducting 2H-MoS2 active sensor regions between metal contacts. The semiconducting region was functionalized with monoclonal antibody Fab (fragment antigen binding) fragments derived from whole antibodies complementary to either SARS-CoV-2 S1 spike protein or Influenza A hemagglutinin using a papain digestion to cleave the antibodies at the disulfide hinges. The high affinity for the MoS2 transducer surface with some density of sulfur vacancies for the antibody fragment base promoted chemisorption with antigen binding regions oriented for interaction with the sample. The angiostatin converting enzyme 2 (ACE2) receptor protein for the SARS-CoV-2 spike glycoprotein, was tethered to a hexa-histidine (his6) tag at its c-terminus both for purification purposes, as well as a motif for binding to MoS2. This modified protein was also investigated as a bio-recognition element. Electrical resistance measurements of sensors functionalized with antibody fragments and exposed to antigen concentrations ranging from 2-20,000 picograms per milliliter revealed selective responses in the presence of complementary antigens with sensitivity to SARS-CoV-2 or influenza A on the order of pg/mL and comparable to gold-standard diagnostics such as Polymerase Chain Reaction (PCR) analysis. Lack of antigen sensitivity for the larger ACE2 BRE further demonstrates the utility of the engineered antibody fragment/transducer interface in bringing the target antigen closer to the transducer surface for sensitivity required for early detection viral diagnostics.

scholarly journals Crystal structure of a glycosylated Fab from an IgM cryoglobulin with properties of a natural proteolytic antibody

2006 ◽  
Vol 395 (3) ◽  
pp. 473-481 ◽  
Author(s):  
Paul A. Ramsland ◽  
Simon S. Terzyan ◽  
Gwendolyn Cloud ◽  
Christina R. Bourne ◽  
William Farrugia ◽  
...  
Keyword(s):  
Serine Proteases ◽  
Three Dimensional ◽  
Catalytic Triad ◽  
Dimensional Structure ◽  
Gel Point ◽  
Antigen Binding ◽  
Antigen Binding Site ◽  
Fab Fragment ◽  
Peripheral Tissues ◽  
Fragment Antigen Binding

The 2.6 Å (1 Å=0.1 nm) resolution structure has been determined for the glycosylated Fab (fragment antigen binding) of an IgM (Yvo) obtained from a subject with Waldenström's macroglobulinaemia. Dynamic light scattering was used to estimate the gel point and monitor the formation of an ordered hydroscopic gel of Yvo IgM upon cooling. If a cryoglobulin forms gels in peripheral tissues and organs, the associated swelling and damage to microvasculature can result in considerable morbidity and mortality. The three-dimensional structure of the branched N-linked oligosaccharide associated with the CH1 domain (first constant domain of heavy chain) is reported. The carbohydrate may act to shield part of the lateral surface of the CH1 domain and crowd the junction between the CH1 and CH2 domains, thereby limiting the segmental flexibility of the Fab arms in intact Yvo IgM, especially at low temperatures. Recently, Yvo IgM was shown to have the properties of a naturally occurring proteolytic antibody [Paul, Karle, Planque, Taguchi, Salas, Nishiyama, Handy, Hunter, Edmundson and Hanson (2004) J. Biol. Chem. 279, 39611–39619; Planque, Bangale, Song, Karle, Taguchi, Poindexter, Bick, Edmundson, Nishiyama and Paul (2004) J. Biol Chem. 279, 14024–14032]. The Yvo protein displayed the ability to cleave, by a nucleophilic mechanism, the amide bonds of a variety of serine protease substrates and the gp120 coat protein of HIV. An atypical serine, arginine and glutamate motif is located in the middle of the Yvo antigen-binding site and displays an overall geometry that mimics the classical serine, histidine and aspartate catalytic triad of serine proteases. Our present findings indicate that pre-existing or natural antibodies can utilize at least one novel strategy for the cleavage of peptide bonds.

scholarly journals Role of husband’s attitude towards the usage of contraceptives for unmet need of family planning among married women of reproductive age in Pakistan

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Muhammad Farhan Asif ◽  
Zahid Pervaiz ◽  
Jawad Rahim Afridi ◽  
Ghulam Abid ◽  
Zohra S. Lassi
Keyword(s):  
Family Planning ◽  
Unmet Need ◽  
Wealth Quintile ◽  
Married Women ◽  
Reproductive Age ◽  
Women Of Reproductive Age ◽  
Family Planning Programs ◽  
Contraception Use ◽  
Wide Range

Abstract Background Family planning services deliver a wide range of benefits to the well-being of females and the community. It can curtail the risk of maternal and neonatal mortality through the reduction in abortions and pregnancies. The government of Pakistan has been struggling to convince people about the usefulness of family planning programs. However, different factors related to social norms, values, and culture are important to determine the success of these programs. One such factor is the patriarchal structure of Pakistani society where most of the household decisions are made by men. The objective of this research is to examine the role of the husband’s attitude towards the usage of contraceptives for the unmet need of family planning (UMNFP) among married women of reproductive age (MWRA) in Pakistan. Method The dataset of Pakistan Demographic and Health Survey 2017–18 is utilized to examine the role of the husband’s attitude towards the usage of contraceptives in UMNFP among MWRA in Pakistan. Results The UMNFP was considerably lower among MWRA between 40 years and above compared to women 15–19 years. The odds of UMNFP were higher among women and men who were educated up to the primary level compared to those with no education. Odds of UMNFP were higher among women from the poor wealth quintile compared to the poorest wealth quintile; similarly, it was significantly lower among women who were from the richer and the richest wealth quintile compared to the poorest wealth quintile. The odds of UMNFP were lower among women who were employed compared to those who were not employed. Lastly, the odds of UMNFP were higher among women whose husbands opposed to using contraceptives, who perceived that there was a religious prohibition for such use and when a decision on the contraception use was solely made by the husband. Conclusions Husband’s attitude towards the usage of contraceptives is an important predictor of UMNFP. Liaising with the community and religious leaders to persuade people particularly men about the usefulness of family planning programs and encouraging men to understand their women’s say in using contraceptives should be encouraged.

scholarly journals The mycoplasma surface proteins MIB and MIP promote the dissociation of the antibody-antigen interaction

2021 ◽  
Vol 7 (10) ◽  
pp. eabf2403
Author(s):  
Pierre Nottelet ◽  
Laure Bataille ◽  
Geraldine Gourgues ◽  
Robin Anger ◽  
Carole Lartigue ◽  
...  
Keyword(s):  
Surface Proteins ◽  
Mycoplasma Species ◽  
Antigen Binding ◽  
Antigen Binding Site ◽  
Fab Fragment ◽  
Cryo Electron Microscopy ◽  
Antigen Complex ◽  
Antigen Interaction ◽  
Immunoglobulin Binding

Mycoplasma immunoglobulin binding (MIB) and mycoplasma immunoglobulin protease (MIP) are surface proteins found in the majority of mycoplasma species, acting sequentially to capture antibodies and cleave off their VH domains. Cryo–electron microscopy structures show how MIB and MIP bind to a Fab fragment in a “hug of death” mechanism. As a result, the orientation of the VL and VH domains is twisted out of alignment, disrupting the antigen binding site. We also show that MIB-MIP has the ability to promote the dissociation of the antibody-antigen complex. This system is functional in cells and protects mycoplasmas from antibody-mediated agglutination. These results highlight the key role of the MIB-MIP system in immunity evasion by mycoplasmas through an unprecedented mechanism, and open exciting perspectives to use these proteins as potential tools in the antibody field.

scholarly journals Double moiré localized plasmon structured illumination microscopy

Nanophotonics ◽  
2020 ◽  
Vol 0 (0) ◽  
Author(s):  
Ruslan Röhrich ◽  
A. Femius Koenderink
Keyword(s):  
Near Field ◽  
Super Resolution ◽  
Reconstruction Algorithm ◽  
Structured Illumination ◽  
Structured Illumination Microscopy ◽  
Experimental Realization ◽  
Spatial Frequencies ◽  
Wide Range ◽  
Localized Plasmon ◽  
Plasmonic Grating

AbstractStructured illumination microscopy (SIM) is a well-established fluorescence imaging technique, which can increase spatial resolution by up to a factor of two. This article reports on a new way to extend the capabilities of structured illumination microscopy, by combining ideas from the fields of illumination engineering and nanophotonics. In this technique, plasmonic arrays of hexagonal symmetry are illuminated by two obliquely incident beams originating from a single laser. The resulting interference between the light grating and plasmonic grating creates a wide range of spatial frequencies above the microscope passband, while still preserving the spatial frequencies of regular SIM. To systematically investigate this technique and to contrast it with regular SIM and localized plasmon SIM, we implement a rigorous simulation procedure, which simulates the near-field illumination of the plasmonic grating and uses it in the subsequent forward imaging model. The inverse problem, of obtaining a super-resolution (SR) image from multiple low-resolution images, is solved using a numerical reconstruction algorithm while the obtained resolution is quantitatively assessed. The results point at the possibility of resolution enhancements beyond regular SIM, which rapidly vanishes with the height above the grating. In an initial experimental realization, the existence of the expected spatial frequencies is shown and the performance of compatible reconstruction approaches is compared. Finally, we discuss the obstacles of experimental implementations that would need to be overcome for artifact-free SR imaging.

Article Commentary: Defining New Directions for More Effective Management of Surgical Pain in the United States: Highlights of the Inaugural Surgical Pain Congress™

2014 ◽  
Vol 80 (3) ◽  
pp. 219-228 ◽  
Author(s):  
Girish P. Joshi ◽  
David E. Beck ◽  
Roger Hill Emerson ◽  
Thomas M. Halaszynski ◽  
Jonathan S. Jahr ◽  
...  
Keyword(s):  
Patient Satisfaction ◽  
Pain Management ◽  
Best Practice ◽  
Task Force ◽  
Unmet Need ◽  
The United States ◽  
Cost Of Care ◽  
Inaugural Meeting ◽  
Surgical Pain ◽  
Wide Range

Despite advances in pharmacologic options for the management of surgical pain, there appears to have been little or no overall improvement over the last two decades in the level of pain experienced by patients. The importance of adequate and effective surgical pain management, however, is clear, because inadequate pain control 1) has a wide range of undesirable physiologic and immunologic effects; 2) is associated with poor surgical outcomes; 3) has increased probability of readmission; and 4) adversely affects the overall cost of care as well as patient satisfaction. There is a clear unmet need for a national surgical pain management consensus task force to raise awareness and develop best practice guidelines for improving surgical pain management, patient safety, patient satisfaction, rapid postsurgical recovery, and health economic outcomes. To comprehensively address this need, the multidisciplinary Surgical Pain Congress™ has been established. The inaugural meeting of this Congress (March 8 to 10, 2013, Celebration, Florida) evaluated the current surgical pain management paradigm and identified key components of best practices.

A novel 3D Zn-coordination polymer based on a multiresponsive fluorescent sensor demonstrating outstanding sensitivities and selectivities for the efficient detection of multiple analytes

2021 ◽  
Author(s):  
Yu Liu ◽  
Yan Wang ◽  
Xiao-Sa Zhang ◽  
Yu-Shu Sheng ◽  
Wen-Ze Li ◽  
...  
Keyword(s):  
Detection Limit ◽  
Coordination Polymer ◽  
Fluorescent Sensor ◽  
Lower Detection Limit ◽  
Ph Values ◽  
Efficient Detection ◽  
Wide Range ◽  
Lower Detection ◽  
Multiple Analytes

A Zn-CP exhibits remarkable fluorescence behaviours and stability in a wide range of pH values. It can become an outstanding candidate in the selective sensing of Fe3+, Mg2+, Cr2O72−, MnO4−, NB and NM at a lower detection limit.

scholarly journals Mosaic Super-resolution via Sequential Feature Pyramid Networks

2019 ◽  
Author(s):  
Mehrdad Shoeiby ◽  
Mohammad Ali Armin ◽  
Sadegh Aliakbarian ◽  
Saeed Anwar ◽  
Lars petersson
Keyword(s):  
State Of The Art ◽  
Super Resolution ◽  
Autonomous Driving ◽  
Single Shot ◽  
Current State ◽  
Wide Range ◽  
Feature Pyramid ◽  
Novel Method ◽  
Convolutional Lstm ◽  
Mosaic Images

<div>Advances in the design of multi-spectral cameras have</div><div>led to great interests in a wide range of applications, from</div><div>astronomy to autonomous driving. However, such cameras</div><div>inherently suffer from a trade-off between the spatial and</div><div>spectral resolution. In this paper, we propose to address</div><div>this limitation by introducing a novel method to carry out</div><div>super-resolution on raw mosaic images, multi-spectral or</div><div>RGB Bayer, captured by modern real-time single-shot mo-</div><div>saic sensors. To this end, we design a deep super-resolution</div><div>architecture that benefits from a sequential feature pyramid</div><div>along the depth of the network. This, in fact, is achieved</div><div>by utilizing a convolutional LSTM (ConvLSTM) to learn the</div><div>inter-dependencies between features at different receptive</div><div>fields. Additionally, by investigating the effect of different</div><div>attention mechanisms in our framework, we show that a</div><div>ConvLSTM inspired module is able to provide superior at-</div><div>tention in our context. Our extensive experiments and anal-</div><div>yses evidence that our approach yields significant super-</div><div>resolution quality, outperforming current state-of-the-art</div><div>mosaic super-resolution methods on both Bayer and multi-</div><div>spectral images. Additionally, to the best of our knowledge,</div><div>our method is the first specialized method to super-resolve</div><div>mosaic images, whether it be multi-spectral or Bayer.</div><div><br></div>

scholarly journals Characterization and validation of fragment antigen-binding (Fab) antibody-based immunoassay for deoxymiroestrol, a potent phytoestrogen from Pueraria candollei

2021 ◽  
Author(s):  
Worapol Sae-foo ◽  
Supaluk Krittanai ◽  
Wipawee Juengsanguanpornsuk ◽  
Gorawit Yusakul ◽  
Tharita Kitisripanya ◽  
...  
Keyword(s):  
Quantitative Method ◽  
Estrogenic Activity ◽  
Limit Of Detection ◽  
Hormone Replacement ◽  
Enzyme Linked Immunosorbent Assay ◽  
Antigen Binding ◽  
Pueraria Candollei ◽  
Specific Determination ◽  
Fragment Antigen Binding

Deoxymiroestrol is the most potent phytoestrogen in chromenes group that has been found in Pueraria candollei, Thai name known as Kwao Krua Khao. Several studies reported estrogenic activity of P. candollei in order to using as hormone replacement therapy for postmenopausal women. Previously, specific determination of deoxymiroestrol content by enzyme-linked immunosorbent assay (ELISA) using polyclonal antibody (pAb) have been reported. However, production of pAb has limitation and variability from different batches. Therefore, in this study, we established quantitative method for determination of deoxymiroestrol using fragment antigen-binding (Fab) antibody-based immunoassay. The developed immunoassay has specificity to deoxymiroestrol with a calibration range of 15.6-1000 ng mL-1. Precision including intra-assay and inter-assay are 1.48-7.11 and 0.58-9.31%, respectively. Accuracy of the assay showed in recovery between 99.77-101.61% when spike deoxymiroestrol standard into the samples. The limit of detection (LOD) is 30.80 ng mL-1. Comparation antibody-based immunoassay for determination of deoxymiroestrol using Fab with pAb was represented consistency (R2 = 0.9807) when analysis roots bark of Pueraria candollei from difference areas. Therefore, this development assay can apply to determine deoxymiroestrol content in the plant samples.

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