Trans-Epithelial Exposure Model (TEEM) - Epithelial Cells and Fibroblasts

2020 ◽  
Author(s):  
Shaun D. McCullough
Keyword(s):  
Particulate Matter ◽  
Epithelial Cells ◽  
Cell Line ◽  
Environmental Protection Agency ◽  
Airway Epithelium ◽  
Exposure Model ◽  
Protection Agency ◽  
The Us ◽  
Trade Names

Abstract This exposure model is meant to serve as an in vitro representation of trans-epithelial effects of airway diesel and particulate matter exposures on fibroblasts in the stroma. The epithelial cells in the airway epithelium are represented by 16HBE cells and the fibroblasts are represented by either the IMR90 cell line or primary fibroblasts.Disclaimer: The contents of this article have been reviewed by the US Environmental Protection Agency and approved for publication and do not necessarily represent Agency policy. Mention of trade names or commercial products does not constitute endorsement or recommendations for use.

scholarly journals Culture of Primary Human Tracheobronchial Epithelial Cells

2021 ◽  
Author(s):  
Lisa Dailey ◽  
Shaun D. McCullough
Keyword(s):  
Epithelial Cells ◽  
Environmental Protection ◽  
Environmental Protection Agency ◽  
Protection Agency ◽  
Commercial Products ◽  
Brush Biopsy ◽  
The Us ◽  
Trade Names ◽  
Agency Policy

Abstract This protocol is intended for culture of primary human tracheobronchial epithelial cells (pHBEC) obtained by brush biopsy during clinical bronchoscopy, or purchased commercially, using Lonza-based medium.Note: This is a historical protocol. At the time of publication, this protocol has been superseded by a different version in the McCullough lab; however, it is being published to support the transparency and reproducibility of other studies by which it is referenced.Disclaimer: The information presented here has been reviewed and approved for publication by the US Environmental Protection Agency do not necessarily represent Agency policy. Mention of trade names or commercial products does not constitute endorsement or recommendations for use.

scholarly journals Culture and Differentiation of Primary Human Tracheobronchial Epithelial Cells Using STEMCELL Technologies Pneumacult Media

2021 ◽  
Author(s):  
Lisa A. Dailey ◽  
Shaun D. McCullough
Keyword(s):  
Epithelial Cells ◽  
Environmental Protection ◽  
Environmental Protection Agency ◽  
Protection Agency ◽  
Commercial Products ◽  
Brush Biopsy ◽  
Previous Version ◽  
The Us ◽  
Scope Of Application ◽  
Trade Names

Abstract NOTE: A PDF methods document is attached in the supplementary materials.SCOPE OF APPLICATION (LIMITATIONS)This method describes the culture and differentiation of primary human tracheobronchial epithelial cells (pHBEC). Cells used for this method can be obtained by brush biopsy during clinical bronchoscopy or purchased commercially. This method replaces the previous version described in (Dailey and McCullough, 2021a; b). Disclaimer: The contents of this article have been reviewed by the US Environmental Protection Agency and approved for publication and do not necessarily represent Agency policy. Mention of trade names or commercial products does not constitute endorsement or recommendations for use.

Isolation of Total RNA with the Life Technologies PureLink Kit

2020 ◽  
Author(s):  
Nicole A. McNabb ◽  
Shaun D. McCullough
Keyword(s):  
Environmental Protection ◽  
Environmental Protection Agency ◽  
Rna Isolation ◽  
Exposure Model ◽  
Protection Agency ◽  
Isolation And Purification ◽  
Total Rna ◽  
The Us ◽  
Trade Names ◽  
Formal Version

Abstract This protocol describes the use of the Life Technologies PureLink RNA isolation kit for the isolation and purification of total RNA from cells grown in culture. Notes describing specific modifications for use with the Trans-Epithelial Exposure Model are included.The attached methods document is a formal version of the information included hereDisclaimer: The contents of this article have been reviewed by the US Environmental Protection Agency and approved for publication and do not necessarily represent Agency policy. Mention of trade names or commercial products does not constitute endorsement or recommendations for use.

Blind trials evaluating in vitro infectivity ofCryptosporidiumoocysts using cell culture immunofluorescence

2007 ◽  
Vol 53 (5) ◽  
pp. 656-663 ◽  
Author(s):  
Zia Bukhari ◽  
David M. Holt ◽  
Michael W. Ware ◽  
Frank W. Schaefer
Keyword(s):  
Cell Culture ◽  
Environmental Protection ◽  
Environmental Protection Agency ◽  
Cost Effective ◽  
Protection Agency ◽  
The Us ◽  
Cryptosporidium Parvum Oocysts ◽  
High Degree ◽  
Blind Trials

An optimized cell culture immunofluorescence (IFA) procedure, using the HCT-8 cell line, was evaluated in blind trials to determine the sensitivity and reproducibility of measuring the infectivity of flow-cytometry-prepared inocula of Cryptosporidium parvum oocysts. In separate trials, suspensions consisting of between 0% and 100% viable oocysts were prepared at the US Environmental Protection Agency, shipped to the American Water Laboratory, and analyzed blindly by cell culture IFA. Data indicated the control (100% live) oocyst suspensions yielded statistically similar results to cell culture dose–response curve data developed previously at the American Water Laboratory. For test samples containing oocyst suspensions of unknown infectivity, cell culture IFA analyses indicated a high degree of correlation (r2= 0.89; n = 26) with the values expected by the US Environmental Protection Agency. Cell culture infectivity correlates well with neonatal mouse infectivity assays, and these blind validation trials provide credibility for the cell culture IFA procedure as a cost-effective and expedient alternative to mouse infectivity assays for determining in vitro infectivity of C. parvum oocysts.

LIVE/DEAD™ Fixable Near-IR Cell Viability Assay

2020 ◽  
Author(s):  
Samantha C. Faber ◽  
Shaun McCullough
Keyword(s):  
Cell Viability ◽  
Environmental Protection Agency ◽  
Protection Agency ◽  
Cell Viability Assay ◽  
Antibody Staining ◽  
Viability Assay ◽  
Near Ir ◽  
The Us ◽  
Trade Names ◽  
Formal Version

Abstract This protocol is used to determine the viability of cells prior to the fixation and permeabilization required for intracellular antibody staining or prior to elimination of biohazardous materials using formaldehyde fixation. The attached methods document is a formal version of the information included here.Disclaimer: The contents of this article have been reviewed by the US Environmental Protection Agency and approved for publication and do not necessarily represent Agency policy. Mention of trade names or commercial products does not constitute endorsement or recommendations for use.

Measurement of Trans-Epithelial Electrical Resistance with EVOM2 and Chopstick Electrode

2020 ◽  
Author(s):  
Shaun D. McCullough
Keyword(s):  
Environmental Protection ◽  
Environmental Protection Agency ◽  
Electrical Resistance ◽  
Cell Monolayer ◽  
Protection Agency ◽  
Commercial Products ◽  
The Us ◽  
Trade Names ◽  
Agency Policy

Abstract Trans-epithelial Electrical Resistance (TEER) can be used as a measure of cell monolayer confluence, health, and integrity. An Epithelial Voltohmmeter (EVOM) is used to take TEER measurements. This method details how to take TEER readings using an EVOM with chopstick electrode.Please note that chopstick electrodes are only designed for use with 12 mm inserts.Disclaimer: The contents of this article have been reviewed by the US Environmental Protection Agency and approved for publication and do not necessarily represent Agency policy. Mention of trade names or commercial products does not constitute endorsement or recommendations for use.

scholarly journals Culture of IMR90 Cells in Advanced MEM with Reduced Serum

2021 ◽  
Author(s):  
Nicholas M. Mallek ◽  
Shaun D. McCullough
Keyword(s):  
Environmental Protection Agency ◽  
Human Lung ◽  
Fibroblast Cell ◽  
Lung Fibroblast ◽  
Protection Agency ◽  
Human Lung Fibroblast ◽  
Lung Fibroblast Cell Line ◽  
The Us ◽  
Scope Of Application ◽  
Trade Names

Abstract NOTE: Methods document and worksheet versions of this method are attached as PDFs.SCOPE OF APPLICATION (LIMITATIONS)This method describes the thawing, culturing, and cryopreservation of the human lung fibroblast cell line IMR90 in Advanced MEM-based growth medium. Disclaimer: The contents of this article have been reviewed by the US Environmental Protection Agency and approved for publication and do not necessarily represent Agency policy. Mention of trade names or commercial products does not constitute endorsement or recommendations for use.

Production and Concentration of Recombinant Lentiviral Particles

2020 ◽  
Author(s):  
Shaun D. McCullough
Keyword(s):  
Environmental Protection ◽  
Environmental Protection Agency ◽  
Protection Agency ◽  
Commercial Products ◽  
The Us ◽  
Trade Names ◽  
Formal Version ◽  
Agency Policy

Abstract This protocol describes the production of recombinant lentivirus by transfection of HEK293T cells with lentiviral packaging vectors and subsequent concentration by ultracentrifugation. The attached methods document is a formal version of the information included here.Disclaimer: The contents of this article have been reviewed by the US Environmental Protection Agency and approved for publication and do not necessarily represent Agency policy. Mention of trade names or commercial products does not constitute endorsement or recommendations for use.

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