Title

Detection of Leishmania Parasites via Flow Cytometry

Presenter Information

Analiese Wenger

Document Type

Oral Presentation

Location

SURC 140

Start Date

21-5-2015

End Date

21-5-2015

Keywords

Leishmania, Flow Cytometry, Macrophage Infection

Abstract

Leishmania species cause a spectrum of diseases collectively referred to as the Leishmaniases. These unicellular parasites generate 1.3 million new cases of infection worldwide as well as cause an approximated 20,000 to 30,000 deaths annually. Current techniques for studying Leishmaniasis in laboratory animals require laborious serial plating of dilution series, incubations of samples for weeks, followed by microscopic screening of each individual well in 96 well plates. Through the generous funding of both the W. M. Keck Foundation and the Murdock Fund, Biological Sciences recently obtained a flow cytometer. This study aimed at assessing if flow cytometry could differentiate cells infected with Leishmania parasites from uninfected cells. Two species of the parasite, L. infantum and L. major, were stained with an antibody conjugated to a florescent fluorochrome, using both flow cytometry and photomicroscopy to detect the parasites. Fluorescent staining of parasites illustrated a shift in florescence intensity using flow cytometry and easily visualized parasites using microscopy. However, a modified protocol for intracellular staining of the murine macrophage cell line, J774A.1, did not reveal a separation between the infected cells and uninfected cells using flow cytometry. The parasites within cells were also undetectable using fluorescent microscopy. Although the hypothesis that there would be a detectable segregation between infected and uninfected cells was not met, this work will continue to attempt to develop what would be a time saving tool in Leishmania research.

Faculty Mentor(s)

Gabrielle Stryker, Blaise Dondji

Department/Program

Biological Sciences

Additional Mentoring Department

Biological Sciences

Additional Mentoring Department

Biological Sciences

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May 21st, 3:00 PM May 21st, 3:20 PM

Detection of Leishmania Parasites via Flow Cytometry

SURC 140

Leishmania species cause a spectrum of diseases collectively referred to as the Leishmaniases. These unicellular parasites generate 1.3 million new cases of infection worldwide as well as cause an approximated 20,000 to 30,000 deaths annually. Current techniques for studying Leishmaniasis in laboratory animals require laborious serial plating of dilution series, incubations of samples for weeks, followed by microscopic screening of each individual well in 96 well plates. Through the generous funding of both the W. M. Keck Foundation and the Murdock Fund, Biological Sciences recently obtained a flow cytometer. This study aimed at assessing if flow cytometry could differentiate cells infected with Leishmania parasites from uninfected cells. Two species of the parasite, L. infantum and L. major, were stained with an antibody conjugated to a florescent fluorochrome, using both flow cytometry and photomicroscopy to detect the parasites. Fluorescent staining of parasites illustrated a shift in florescence intensity using flow cytometry and easily visualized parasites using microscopy. However, a modified protocol for intracellular staining of the murine macrophage cell line, J774A.1, did not reveal a separation between the infected cells and uninfected cells using flow cytometry. The parasites within cells were also undetectable using fluorescent microscopy. Although the hypothesis that there would be a detectable segregation between infected and uninfected cells was not met, this work will continue to attempt to develop what would be a time saving tool in Leishmania research.