scholarly journals Estimation of Saprolite Thickness Needed to Remove E. coli from Wastewater

2021 ◽  
Vol 11 (5) ◽  
pp. 2066
Author(s):  
Michael J. Vepraskas ◽  
Aziz Amoozegar ◽  
Terrence Gardner
Keyword(s):  
North Carolina ◽  
Water Retention ◽  
Coarse Sand ◽  
Retention Data ◽  
Clay Loam ◽  
Septic System ◽  
Simple Method ◽  
E Coli ◽  
Biological Contaminants

Saprolite, weathered bedrock, is being used to dispose of domestic sewage through septic system drainfields, but the thickness of saprolite needed to remove biological contaminants is unknown for most saprolites. This study developed and tested a simple method for estimating the thickness of saprolite needed below septic drainlines to filter E. coli from wastewater using estimates of the volume of pores that are smaller than the length of the coliform (≤10 μm). Particle size distribution (texture) and water retention data were obtained for 12 different saprolites from the Piedmont and Mountain regions of North Carolina (N.C.). Saprolite textures ranged from clay loam to coarse sand. The volume of pores with diameters ≤10 μm were determined by water retention measurements for each saprolite. The data were used in an equation to estimate the saprolite thickness needed to filter E. coli. The estimated saprolite thicknesses ranged from 36 cm in the clay loam to 113 cm for the coarse sand. The average thickness across all samples was 58 cm. Saprolite thickness estimates increased as silt percentage decreased and as sand percentage and in situ saturated hydraulic conductivity increased. Silt percentage may be most useful for estimating appropriate saprolite thicknesses in the field.

Rotary Beveling for In Situ Thin-Section Microscopy of Cell Cultures

1981 ◽  
Vol 39 ◽  
pp. 550-551
Author(s):  
Dean A. Handley ◽  
Jack T. Alexander ◽  
Shu Chien
Keyword(s):  
Cell Growth ◽  
Cell Cultures ◽  
Molecular Interactions ◽  
Convenient Method ◽  
Petri Dish ◽  
Simple Method ◽  
Thin Section Microscopy ◽  
Thin Sectioning ◽  
Organic Fluids

In situ preparation of cell cultures for ultrastructural investigations is a convenient method by which fixation, dehydration and embedment are carried out in the culture petri dish. The in situ method offers the advantage of preserving the native orientation of cell-cell interactions, junctional regions and overlapping configurations. In order to section after embedment, the petri dish is usually separated from the polymerized resin by either differential cryo-contraction or solvation in organic fluids. The remaining resin block must be re-embedded before sectioning. Although removal of the petri dish may not disrupt the native cellular geometry, it does sacrifice what is now recognized as an important characteristic of cell growth: cell-substratum molecular interactions. To preserve the topographic cell-substratum relationship, we developed a simple method of tapered rotary beveling to reduce the petri dish thickness to a dimension suitable for direct thin sectioning.

Optics-Free Visualization of Proteins in Single Cells by Time of Flight-Secondary Ion Mass Spectrometry Coupled with Genetically Encoded Chemical Tags

2020 ◽  
Author(s):  
Feifei Jia ◽  
Jie Wang ◽  
Yanyan Zhang ◽  
Qun Luo ◽  
Luyu Qi ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Secondary Ion Mass Spectrometry ◽  
Single Cells ◽  
Time Of Flight ◽  
E Coli ◽  
Tof Sims ◽  
Ion Mass Spectrometry ◽  
Chemical Tags ◽  
Secondary Ion

<p></p><p><i>In situ</i> visualization of proteins of interest at single cell level is attractive in cell biology, molecular biology and biomedicine, which usually involves photon, electron or X-ray based imaging methods. Herein, we report an optics-free strategy that images a specific protein in single cells by time of flight-secondary ion mass spectrometry (ToF-SIMS) following genetic incorporation of fluorine-containing unnatural amino acids as a chemical tag into the protein via genetic code expansion technique. The method was developed and validated by imaging GFP in E. coli and human HeLa cancer cells, and then utilized to visualize the distribution of chemotaxis protein CheA in E. coli cells and the interaction between high mobility group box 1 protein and cisplatin damaged DNA in HeLa cells. The present work highlights the power of ToF-SIMS imaging combined with genetically encoded chemical tags for <i>in situ </i>visualization of proteins of interest as well as the interactions between proteins and drugs or drug damaged DNA in single cells.</p><p></p>

A Mild and Simple Method for Making MIDA Boronates

2020 ◽  
Author(s):  
Aidan Kelly ◽  
Peng-Jui (Ruby) Chen ◽  
Jenna Klubnick ◽  
Daniel J. Blair ◽  
Martin D. Burke
Keyword(s):  
Organic Synthesis ◽  
Boronic Acids ◽  
Simple Method ◽  
Direct Preparation ◽  
Synthesis Procedure ◽  
Harsh Conditions

<div> <div> <div> <p>Existing methods for making MIDA boronates require harsh conditions and complex procedures to achieve dehydration. Here we disclose that a pre-dried form of MIDA, MIDA anhydride, acts as both a source of the MIDA ligand and an in situ desiccant to enable a mild and simple MIDA boronate synthesis procedure. This method expands the range of sensitive boronic acids that can be converted into their MIDA boronate counterparts. Further utilizing unique properties of MIDA boronates, we have developed a MIDA Boronate Maker Kit which enables the direct preparation and purification of MIDA boronates from boronic acids using only heating and centrifuge equipment that is widely available in labs that do not specialize in organic synthesis. </p> </div> </div> </div>

scholarly journals DnaA, the Initiator of Escherichia coliChromosomal Replication, Is Located at the Cell Membrane

2000 ◽  
Vol 182 (9) ◽  
pp. 2604-2610 ◽  
Author(s):  
Gillian Newman ◽  
Elliott Crooke
Keyword(s):  
Cell Membrane ◽  
Spatial Organization ◽  
Active Form ◽  
Chromosomal Replication ◽  
E Coli ◽  
Dnaa Protein ◽  
Section Electron ◽  
Acidic Phospholipids

ABSTRACT Given the lack of a nucleus in prokaryotic cells, the significance of spatial organization in bacterial chromosome replication is only beginning to be fully appreciated. DnaA protein, the initiator of chromosomal replication in Escherichia coli, is purified as a soluble protein, and in vitro it efficiently initiates replication of minichromosomes in membrane-free DNA synthesis reactions. However, its conversion from a replicatively inactive to an active form in vitro occurs through its association with acidic phospholipids in a lipid bilayer. To determine whether the in situ residence of DnaA protein is cytoplasmic, membrane associated, or both, we examined the cellular location of DnaA using immunogold cryothin-section electron microscopy and immunofluorescence. Both of these methods revealed that DnaA is localized at the cell membrane, further suggesting that initiation of chromosomal replication in E. coli is a membrane-affiliated event.

scholarly journals Assessing an Atmospheric Correction Algorithm for Time Series of Satellite-Based Water-Leaving Reflectance Using Match-Up Sites in Australian Coastal Waters

2021 ◽  
Vol 13 (10) ◽  
pp. 1927
Author(s):  
Fuqin Li ◽  
David Jupp ◽  
Thomas Schroeder ◽  
Stephen Sagar ◽  
Joshua Sixsmith ◽  
...  
Keyword(s):  
Coastal Waters ◽  
Atmospheric Correction ◽  
Correction Method ◽  
In Situ Measurements ◽  
Landsat 8 ◽  
Correction Algorithm ◽  
Simple Method ◽  
North East ◽  
The Impact

An atmospheric correction algorithm for medium-resolution satellite data over general water surfaces (open/coastal, estuarine and inland waters) has been assessed in Australian coastal waters. In situ measurements at four match-up sites were used with 21 Landsat 8 images acquired between 2014 and 2017. Three aerosol sources (AERONET, MODIS ocean aerosol and climatology) were used to test the impact of the selection of aerosol optical depth (AOD) and Ångström coefficient on the retrieved accuracy. The initial results showed that the satellite-derived water-leaving reflectance can have good agreement with the in situ measurements, provided that the sun glint is handled effectively. Although the AERONET aerosol data performed best, the contemporary satellite-derived aerosol information from MODIS or an aerosol climatology could also be as effective, and should be assessed with further in situ measurements. Two sun glint correction strategies were assessed for their ability to remove the glint bias. The most successful one used the average of two shortwave infrared (SWIR) bands to represent sun glint and subtracted it from each band. Using this sun glint correction method, the mean all-band error of the retrieved water-leaving reflectance at the Lucinda Jetty Coastal Observatory (LJCO) in north east Australia was close to 4% and unbiased over 14 acquisitions. A persistent bias in the other strategy was likely due to the sky radiance being non-uniform for the selected images. In regard to future options for an operational sun glint correction, the simple method may be sufficient for clear skies until a physically based method has been established.

scholarly journals Microchannelled alkylated chitosan sponge to treat noncompressible hemorrhages and facilitate wound healing

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Xinchen Du ◽  
Le Wu ◽  
Hongyu Yan ◽  
Zhuyan Jiang ◽  
Shilin Li ◽  
...  
Keyword(s):  
Wound Healing ◽  
Shape Recovery ◽  
Parenchymal Cell ◽  
Gelatin Sponge ◽  
Defect Model ◽  
Liver Parenchymal Cell ◽  
E Coli ◽  
Tissue Integration ◽  
3D Printed

AbstractDeveloping an anti-infective shape-memory hemostatic sponge able to guide in situ tissue regeneration for noncompressible hemorrhages in civilian and battlefield settings remains a challenge. Here we engineer hemostatic chitosan sponges with highly interconnective microchannels by combining 3D printed microfiber leaching, freeze-drying, and superficial active modification. We demonstrate that the microchannelled alkylated chitosan sponge (MACS) exhibits the capacity for water and blood absorption, as well as rapid shape recovery. We show that compared to clinically used gauze, gelatin sponge, CELOX™, and CELOX™-gauze, the MACS provides higher pro-coagulant and hemostatic capacities in lethally normal and heparinized rat and pig liver perforation wound models. We demonstrate its anti-infective activity against S. aureus and E. coli and its promotion of liver parenchymal cell infiltration, vascularization, and tissue integration in a rat liver defect model. Overall, the MACS demonstrates promising clinical translational potential in treating lethal noncompressible hemorrhage and facilitating wound healing.

scholarly journals Enzymatic synthesis and phosphorolysis of 4(2)-thioxo- and 6(5)-azapyrimidine nucleosides by E. coli nucleoside phosphorylases

2016 ◽  
Vol 12 ◽  
pp. 2588-2601 ◽  
Author(s):  
Vladimir A Stepchenko ◽  
Anatoly I Miroshnikov ◽  
Frank Seela ◽  
Igor A Mikhailopulo
Keyword(s):  
Purine Nucleoside Phosphorylase ◽  
Reaction Conditions ◽  
E Coli ◽  
No Formation ◽  
Structural Peculiarities ◽  
Substrate Properties ◽  
Nucleoside Phosphorylases ◽  
Derivatives Of

The trans-2-deoxyribosylation of 4-thiouracil (4SUra) and 2-thiouracil (2SUra), as well as 6-azauracil, 6-azathymine and 6-aza-2-thiothymine was studied using dG and E. coli purine nucleoside phosphorylase (PNP) for the in situ generation of 2-deoxy-α-D-ribofuranose-1-phosphate (dRib-1P) followed by its coupling with the bases catalyzed by either E. coli thymidine (TP) or uridine (UP) phosphorylases. 4SUra revealed satisfactory substrate activity for UP and, unexpectedly, complete inertness for TP; no formation of 2’-deoxy-2-thiouridine (2SUd) was observed under analogous reaction conditions in the presence of UP and TP. On the contrary, 2SU, 2SUd, 4STd and 2STd are good substrates for both UP and TP; moreover, 2SU, 4STd and 2’-deoxy-5-azacytidine (Decitabine) are substrates for PNP and the phosphorolysis of the latter is reversible. Condensation of 2SUra and 5-azacytosine with dRib-1P (Ba salt) catalyzed by the accordant UP and PNP in Tris∙HCl buffer gave 2SUd and 2’-deoxy-5-azacytidine in 27% and 15% yields, respectively. 6-Azauracil and 6-azathymine showed good substrate properties for both TP and UP, whereas only TP recognizes 2-thio-6-azathymine as a substrate. 5-Phenyl and 5-tert-butyl derivatives of 6-azauracil and its 2-thioxo derivative were tested as substrates for UP and TP, and only 5-phenyl- and 5-tert-butyl-6-azauracils displayed very low substrate activity. The role of structural peculiarities and electronic properties in the substrate recognition by E. coli nucleoside phosphorylases is discussed.

scholarly journals Combining Augmented Radiotherapy and Immunotherapy through a Nano-Gold and Bacterial Outer-Membrane Vesicle Complex for the Treatment of Glioblastoma

Nanomaterials ◽  
2021 ◽  
Vol 11 (7) ◽  
pp. 1661
Author(s):  
Mei-Hsiu Chen ◽  
Tse-Ying Liu ◽  
Yu-Chiao Chen ◽  
Ming-Hong Chen
Keyword(s):  
Outer Membrane ◽  
Membrane Vesicle ◽  
Glioma Cells ◽  
Membrane Vesicles ◽  
Outer Membrane Vesicles ◽  
E Coli ◽  
Tumor Bearing ◽  
Nano Gold ◽  
Gl261 Cell

Glioblastoma, formerly known as glioblastoma multiforme (GBM), is refractory to existing adjuvant chemotherapy and radiotherapy. We successfully synthesized a complex, Au–OMV, with two specific nanoparticles: gold nanoparticles (AuNPs) and outer-membrane vesicles (OMVs) from E. coli. Au–OMV, when combined with radiotherapy, produced radiosensitizing and immuno-modulatory effects that successfully suppressed tumor growth in both subcutaneous G261 tumor-bearing and in situ (brain) tumor-bearing C57BL/6 mice. Longer survival was also noted with in situ tumor-bearing mice treated with Au–OMV and radiotherapy. The mechanisms for the successful treatment were evaluated. Intracellular reactive oxygen species (ROS) greatly increased in response to Au–OMV in combination with radiotherapy in G261 glioma cells. Furthermore, with a co-culture of G261 glioma cells and RAW 264.7 macrophages, we found that GL261 cell viability was related to chemotaxis of macrophages and TNF-α production.

scholarly journals 762 A combination of functional biomarkers improves identification of the tumor-specific reactive T cell repertoire

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A810-A810
Author(s):  
Arianna Draghi ◽  
Katja Harbst ◽  
Inge Svane ◽  
Marco Donia
Keyword(s):  
T Cells ◽  
T Cell ◽  
De Novo ◽  
Autologous Tumor ◽  
T Cell Repertoire ◽  
Cell Repertoire ◽  
Simple Method ◽  
Functional Biomarkers

BackgroundDetecting the entire repertoire of tumor-specific reactive T cells is essential for investigating the broad range of T cell functions in the tumor-microenvironment. At present, assays identifying tumor-specific functional activation measure either upregulation of specific surface molecules, de novo production of the most common antitumor cytokines or mobilization of cytotoxic granules.MethodsIn this study, we combined transcriptomic analyses of tumor-specific reactive tumorinfiltrating lymphocytes (TILs), TIL-autologous tumor cell co-cultures and commonly used established detection protocols to develop an intracellular flow cytometry staining method encompassing simultaneous detection of intracellular CD137, de novo production of TNF and IFNy and extracellular mobilization of CD107a.ResultsThis approach enabled the identification of a larger fraction of tumor-specific reactive T cells in vitro compared to standard methods, revealing the existence of multiple distinct functional clusters of tumor-specific reactive TILs. Publicly available datasets of fresh tumor single-cell RNA-sequencing from four cancer types were investigated to confirm that these functional biomarkers identified distinct functional clusters forming the entire repertoire of tumor-specific reactive T cells in situ.ConclusionsIn conclusion, we describe a simple method using a combination of functional biomarkers that improves identification of the tumor-specific reactive T cell repertoire in vitro and in situ.

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