Methods for In Vivo/Ex Vivo Analysis of Antimicrobial Peptides in Bacterial Keratitis: siRNA Knockdown, Colony Counts, Myeloperoxidase, Immunostaining, and RT-PCR Assays

Satya Sree Kolar; Hasna Baidouri; Maria Luisa Mangoni; Alison McDermott

doi:10.1007/978-1-4939-6737-7_30

Methods for In Vivo/Ex Vivo Analysis of Antimicrobial Peptides in Bacterial Keratitis: siRNA Knockdown, Colony Counts, Myeloperoxidase, Immunostaining, and RT-PCR Assays

Satya Sree Kolar, Hasna Baidouri, Maria Luisa Mangoni, Alison McDermott

Research output: Chapter in Book/Report/Conference proceeding › Chapter › peer-review

Abstract

Antimicrobial peptides (AMPs) are essential components of the innate immune response. They have direct killing ability as well as immunomodulatory functions. Here, we describe techniques to identify specific AMPs involved in the protection against microbial keratitis, a vision threatening infection of the cornea of the eye which is the most serious complication of contact lens wear. Specifically we detail the use of siRNA technology to temporarily knockdown AMP expression at the murine ocular surface in vivo and then describe ex vivo assays to determine the level of bacteria, relative number of neutrophils, and levels of cytokines, chemokines, and AMPs in infected corneas.

Original language	English
Title of host publication	Antimicrobial Peptides
Editors	Paul Hansen
Place of Publication	London
Publisher	Springer
Pages	411-425
Volume	1548
ISBN (Print)	978-1-4939-6735-3
DOIs	https://doi.org/10.1007/978-1-4939-6737-7_30
Publication status	Published - 25 Dec 2016

Publication series

Name	Methods in Molecular Biology
Publisher	Springer
ISSN (Electronic)	1064-3745

Keywords

Antimicrobial peptide
Inflammation
Keratitis
Neutrophil
Pseudomonas

Access to Document

10.1007/978-1-4939-6737-7_30

Cite this

Kolar, S. S., Baidouri, H., Mangoni, M. L., & McDermott, A. (2016). Methods for In Vivo/Ex Vivo Analysis of Antimicrobial Peptides in Bacterial Keratitis: siRNA Knockdown, Colony Counts, Myeloperoxidase, Immunostaining, and RT-PCR Assays. In P. Hansen (Ed.), Antimicrobial Peptides (Vol. 1548, pp. 411-425). (Methods in Molecular Biology). Springer. https://doi.org/10.1007/978-1-4939-6737-7_30

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abstract = "Antimicrobial peptides (AMPs) are essential components of the innate immune response. They have direct killing ability as well as immunomodulatory functions. Here, we describe techniques to identify specific AMPs involved in the protection against microbial keratitis, a vision threatening infection of the cornea of the eye which is the most serious complication of contact lens wear. Specifically we detail the use of siRNA technology to temporarily knockdown AMP expression at the murine ocular surface in vivo and then describe ex vivo assays to determine the level of bacteria, relative number of neutrophils, and levels of cytokines, chemokines, and AMPs in infected corneas.",

keywords = "Antimicrobial peptide, Inflammation, Keratitis, Neutrophil, Pseudomonas",

author = "Kolar, \{Satya Sree\} and Hasna Baidouri and Mangoni, \{Maria Luisa\} and Alison McDermott",

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Kolar, SS, Baidouri, H, Mangoni, ML & McDermott, A 2016, Methods for In Vivo/Ex Vivo Analysis of Antimicrobial Peptides in Bacterial Keratitis: siRNA Knockdown, Colony Counts, Myeloperoxidase, Immunostaining, and RT-PCR Assays. in P Hansen (ed.), Antimicrobial Peptides. vol. 1548, Methods in Molecular Biology, Springer, London, pp. 411-425. https://doi.org/10.1007/978-1-4939-6737-7_30

Methods for In Vivo/Ex Vivo Analysis of Antimicrobial Peptides in Bacterial Keratitis: siRNA Knockdown, Colony Counts, Myeloperoxidase, Immunostaining, and RT-PCR Assays. / Kolar, Satya Sree; Baidouri, Hasna; Mangoni, Maria Luisa et al.
Antimicrobial Peptides. ed. / Paul Hansen. Vol. 1548 London: Springer, 2016. p. 411-425 (Methods in Molecular Biology).

Research output: Chapter in Book/Report/Conference proceeding › Chapter › peer-review

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T1 - Methods for In Vivo/Ex Vivo Analysis of Antimicrobial Peptides in Bacterial Keratitis: siRNA Knockdown, Colony Counts, Myeloperoxidase, Immunostaining, and RT-PCR Assays

AU - Kolar, Satya Sree

AU - Baidouri, Hasna

AU - Mangoni, Maria Luisa

AU - McDermott, Alison

PY - 2016/12/25

Y1 - 2016/12/25

N2 - Antimicrobial peptides (AMPs) are essential components of the innate immune response. They have direct killing ability as well as immunomodulatory functions. Here, we describe techniques to identify specific AMPs involved in the protection against microbial keratitis, a vision threatening infection of the cornea of the eye which is the most serious complication of contact lens wear. Specifically we detail the use of siRNA technology to temporarily knockdown AMP expression at the murine ocular surface in vivo and then describe ex vivo assays to determine the level of bacteria, relative number of neutrophils, and levels of cytokines, chemokines, and AMPs in infected corneas.

AB - Antimicrobial peptides (AMPs) are essential components of the innate immune response. They have direct killing ability as well as immunomodulatory functions. Here, we describe techniques to identify specific AMPs involved in the protection against microbial keratitis, a vision threatening infection of the cornea of the eye which is the most serious complication of contact lens wear. Specifically we detail the use of siRNA technology to temporarily knockdown AMP expression at the murine ocular surface in vivo and then describe ex vivo assays to determine the level of bacteria, relative number of neutrophils, and levels of cytokines, chemokines, and AMPs in infected corneas.

KW - Antimicrobial peptide

KW - Inflammation

KW - Keratitis

KW - Neutrophil

KW - Pseudomonas

UR - https://www.scopus.com/pages/publications/85007482439

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DO - 10.1007/978-1-4939-6737-7_30

M3 - Chapter

SN - 978-1-4939-6735-3

VL - 1548

T3 - Methods in Molecular Biology

SP - 411

EP - 425

BT - Antimicrobial Peptides

A2 - Hansen, Paul

PB - Springer

CY - London

ER -

Kolar SS, Baidouri H, Mangoni ML, McDermott A. Methods for In Vivo/Ex Vivo Analysis of Antimicrobial Peptides in Bacterial Keratitis: siRNA Knockdown, Colony Counts, Myeloperoxidase, Immunostaining, and RT-PCR Assays. In Hansen P, editor, Antimicrobial Peptides. Vol. 1548. London: Springer. 2016. p. 411-425. (Methods in Molecular Biology). doi: 10.1007/978-1-4939-6737-7_30

Methods for In Vivo/Ex Vivo Analysis of Antimicrobial Peptides in Bacterial Keratitis: siRNA Knockdown, Colony Counts, Myeloperoxidase, Immunostaining, and RT-PCR Assays

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