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Standard Bacteriophage Purification Procedures Cause Loss in Numbers and Activity

For decades, bacteriophage purification has followed structured protocols focused on generating high concentrations of phage in manageable volumes. As research moves toward understanding complex phage populations, purification needs have shifted to maximize the amount of phage while maintaining dive...

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Detalles Bibliográficos
Autores principales: Carroll-Portillo, Amanda, Coffman, Cristina N., Varga, Matthew G., Alcock, Joe, Singh, Sudha B., Lin, Henry C.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7924620/
https://www.ncbi.nlm.nih.gov/pubmed/33672780
http://dx.doi.org/10.3390/v13020328
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author Carroll-Portillo, Amanda
Coffman, Cristina N.
Varga, Matthew G.
Alcock, Joe
Singh, Sudha B.
Lin, Henry C.
author_facet Carroll-Portillo, Amanda
Coffman, Cristina N.
Varga, Matthew G.
Alcock, Joe
Singh, Sudha B.
Lin, Henry C.
author_sort Carroll-Portillo, Amanda
collection PubMed
description For decades, bacteriophage purification has followed structured protocols focused on generating high concentrations of phage in manageable volumes. As research moves toward understanding complex phage populations, purification needs have shifted to maximize the amount of phage while maintaining diversity and activity. The effects of standard phage purification procedures such as polyethylene glycol (PEG) precipitation and cesium chloride (CsCl) density gradients on both diversity and activity of a phage population are not known. We have examined the effects of PEG precipitation and CsCl density gradients on a number of known phage (M13, T4, and ΦX 174) of varying structure and size, individually and as mixed sample. Measurement of phage numbers and activity throughout the purification process was performed. We demonstrate that these methods, used routinely to generate “pure” phage samples, are in fact detrimental to retention of phage number and activity; even more so in mixed phage samples. As such, minimal amounts of processing are recommended to introduce less bias and maintain more of a phage population.
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spelling pubmed-79246202021-03-03 Standard Bacteriophage Purification Procedures Cause Loss in Numbers and Activity Carroll-Portillo, Amanda Coffman, Cristina N. Varga, Matthew G. Alcock, Joe Singh, Sudha B. Lin, Henry C. Viruses Article For decades, bacteriophage purification has followed structured protocols focused on generating high concentrations of phage in manageable volumes. As research moves toward understanding complex phage populations, purification needs have shifted to maximize the amount of phage while maintaining diversity and activity. The effects of standard phage purification procedures such as polyethylene glycol (PEG) precipitation and cesium chloride (CsCl) density gradients on both diversity and activity of a phage population are not known. We have examined the effects of PEG precipitation and CsCl density gradients on a number of known phage (M13, T4, and ΦX 174) of varying structure and size, individually and as mixed sample. Measurement of phage numbers and activity throughout the purification process was performed. We demonstrate that these methods, used routinely to generate “pure” phage samples, are in fact detrimental to retention of phage number and activity; even more so in mixed phage samples. As such, minimal amounts of processing are recommended to introduce less bias and maintain more of a phage population. MDPI 2021-02-20 /pmc/articles/PMC7924620/ /pubmed/33672780 http://dx.doi.org/10.3390/v13020328 Text en © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Carroll-Portillo, Amanda
Coffman, Cristina N.
Varga, Matthew G.
Alcock, Joe
Singh, Sudha B.
Lin, Henry C.
Standard Bacteriophage Purification Procedures Cause Loss in Numbers and Activity
title Standard Bacteriophage Purification Procedures Cause Loss in Numbers and Activity
title_full Standard Bacteriophage Purification Procedures Cause Loss in Numbers and Activity
title_fullStr Standard Bacteriophage Purification Procedures Cause Loss in Numbers and Activity
title_full_unstemmed Standard Bacteriophage Purification Procedures Cause Loss in Numbers and Activity
title_short Standard Bacteriophage Purification Procedures Cause Loss in Numbers and Activity
title_sort standard bacteriophage purification procedures cause loss in numbers and activity
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7924620/
https://www.ncbi.nlm.nih.gov/pubmed/33672780
http://dx.doi.org/10.3390/v13020328
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