Poster or Presentation Title

The Importance of Optimal Conditions and Technique in the Culture of RAW 264.7 Murine Macrophages

Presenter Information

Natalie ReighFollow

Location

Hall Memorial Ballroom

Access Type

Open Access

Presentation Type

Poster Presentation

Start Date

4-4-2018 12:00 PM

Department

Biomedical Science

Abstract

RAW 264.7 cells are a commonly used cell line. Inconsistencies in cell culture techniques and a tendency to omit reporting aspects of culture methods may have an impact on the ability to compare research results. We examined the effects of two methods of passage, scraping and trypsinization; cell viability; and the effect of growth media on response to lipopolysaccharide (LPS) derived from 055:B5 E. coli. RAW 264.7 cells were grown in DMEM or RPMI 1640 growth media and passage was performed every 4 days using scraping and trypsinization separately. All experiments were performed between passages 5 through 20. Cells were plated and treated with LPS concentrations of 0, 1, 10, and 100 ng/ml for 24 hours. LPS response was quantified by measuring nitrites via a Greiss reaction. No difference in nitrites was seen between scraping and trypsin passage techniques. The nitrite response of the cells grown in DMEM was at least five-fold greater than the cells grown in RPMI 1640. A correlation was seen between LPS response and the experience of the researcher when compared in series, with novice students producing lower responses. This suggests that experience level can have an unforeseen effect on experimental outcomes.

Faculty Mentor

Dr. David O. Freier

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Apr 4th, 12:00 PM

The Importance of Optimal Conditions and Technique in the Culture of RAW 264.7 Murine Macrophages

Hall Memorial Ballroom

RAW 264.7 cells are a commonly used cell line. Inconsistencies in cell culture techniques and a tendency to omit reporting aspects of culture methods may have an impact on the ability to compare research results. We examined the effects of two methods of passage, scraping and trypsinization; cell viability; and the effect of growth media on response to lipopolysaccharide (LPS) derived from 055:B5 E. coli. RAW 264.7 cells were grown in DMEM or RPMI 1640 growth media and passage was performed every 4 days using scraping and trypsinization separately. All experiments were performed between passages 5 through 20. Cells were plated and treated with LPS concentrations of 0, 1, 10, and 100 ng/ml for 24 hours. LPS response was quantified by measuring nitrites via a Greiss reaction. No difference in nitrites was seen between scraping and trypsin passage techniques. The nitrite response of the cells grown in DMEM was at least five-fold greater than the cells grown in RPMI 1640. A correlation was seen between LPS response and the experience of the researcher when compared in series, with novice students producing lower responses. This suggests that experience level can have an unforeseen effect on experimental outcomes.