Skip to Main Content
 

Global Search Box

 
 
 
 

Files

File List


masters thesis 4-6-16.pdf (2.55 MB)

ETD Abstract Container

Abstract Header

Characterization of Sialic Acid Receptors on MDCK Cells Maintained Under Different Media Conditions by Flow Cytometric Analysis and Implications for Detection of Influenza A Virus.

Nelson, Sarah W
http://orcid.org/0000-0003-3583-450X
http://rave.ohiolink.edu/etdc/view?acc_num=osu1460020380

Abstract Details

2016, Master of Science, Ohio State University, Comparative and Veterinary Medicine.
Abstract Thousands of people become infected with seasonally circulating influenza A virus (IAV) each year and outbreaks of influenza in commercial poultry and swine operations lead to heavy economic losses. In order to subtype and perform diagnostic experiments on IAVs, samples are commonly cultured in embryonating chicken eggs or Madin-Darby Canine Kidney (MDCK) cells. For the current work, it was hypothesized that media conditions would modulate the distribution of receptors on MDCK cells, resulting in alterations in the amount of virus produced by the culture system. MDCK cells were cultured in minimum essential medium (MEM) supplemented with 10% fetal bovine serum (FBS) or two different commercially available serum free media (SFM) types. Distributions of a-2,6 linked and a-2,3 linked sialic acid cell surface receptors were compared by flow cytometric analysis over time. MEM supplemented with 10% FBS resulted in a cycling between high percentages of cells expressing both a-2,6 linked and a-2,3 linked sialic acids following one passage, then high percentages of cells expressing only a-2,3 linked sialic acids the next passage. The two different SFM brands also altered the sialic acid distributions of MDCK cells. Culture in UltraMDCK SFM resulted in a higher percentage of cells expressing both a-2,6 linked and a-2,3 linked sialic acid receptors over 25 passages. Culture in OptiPro SFM resulted in a more variable distribution of each receptor over 25 passages. To determine if the cells were using media nutrients in a manner that might influence the sialic acid distributions on the cells, MDCK cells were seeded into flasks containing UltraMDCK SFM or MEM supplemented with 10% FBS and cultured for seven days without changing media. Three flasks from each media group were analyzed each day for seven days by flow cytometric analysis. Culture in UltraMDCK SFM caused MDCK cells to express both receptors, while culture in MEM with 10% FBS showed variability in the a-2,6 linked and a-2,3 linked sialic acid receptor expression. In the final experiment, effects of media conditions on the amount of IAV recovered from each culture system were determined. Cells were maintained in UltraMDCK SFM or MEM supplemented with 10% FBS, the a-2,6 linked and a-2,3 linked sialic acid receptor distributions on the cells were determined, and tissue culture infective dose 50% experiments were conducted. Cells were plated at a high density so they would be confluent the next day. MDCK cells maintained in SFM expressed predominantly a-2,6 linked sialic acids, while cells maintained in MEM supplemented with 10% FBS expressed more a-2,3 linked sialic acids. The swine origin IAV isolate grew to similar titers in MDCK cells maintained in both SFM and MEM supplemented with 10% FBS. The avian origin IAV isolate grew to significantly lower titers in MDCK cells maintained in MEM supplemented with 10% FBS when compared to growth in cells maintained in SFM. The effects of culture media on the distributions of sialic acids present on MDCK cells should be studied further to better understand the limitations and effects of IAV isolation in these cells.
Ian Davis (Advisor)
76 p.
Cellular Biology; Public Health; Virology
MDCK; cell culture; flow cytometry; influenza

Recommended Citations

Citations

  • Nelson, S. W. (2016). Characterization of Sialic Acid Receptors on MDCK Cells Maintained Under Different Media Conditions by Flow Cytometric Analysis and Implications for Detection of Influenza A Virus. [Master's thesis, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1460020380

    APA Style (7th edition)

  • Nelson, Sarah. Characterization of Sialic Acid Receptors on MDCK Cells Maintained Under Different Media Conditions by Flow Cytometric Analysis and Implications for Detection of Influenza A Virus. 2016. Ohio State University, Master's thesis. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1460020380.

    MLA Style (8th edition)

  • Nelson, Sarah. "Characterization of Sialic Acid Receptors on MDCK Cells Maintained Under Different Media Conditions by Flow Cytometric Analysis and Implications for Detection of Influenza A Virus." Master's thesis, Ohio State University, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=osu1460020380

    Chicago Manual of Style (17th edition)

osu1460020380
1,407
© 2016, all rights reserved.
This open access ETD is published by The Ohio State University and OhioLINK.