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Team:Warsaw/Calendar-Stage1/23 July 2010
From 2010.igem.org
23.07.2010 Friday
Note: We are not totally satisfied with the results, because of the performance of the cells during the flow cytometry examination, they where adhering to each other.
Previous 2 days: Set up and culturing Hela cell culture [Kamil]
Conducted according to the protocol included in "Quantitative measurement of mamallian cell invasion by bacteria using flow cytometry" standard (RFC 59 standard point 5)
Previous day: Set up liquid cultures of E.coli with different constructs [Ania P.]
Conducted according to the protocol included in "Quantitative measurement of mamallian cell invasion by bacteria using flow cytometry" standard (RFC 59 standard point 5)
Testing the invasivnes of E.coli in Hela cells [Kamil,Cherry]
1. Incubation of Hela cells with E.coli with different constructs.
2. Preparation of samples with incubated cells and examining them in flow cytometer.
Conducted according to the protocol included in "Quantitative measurement of mamallian cell invasion by bacteria using flow cytometry" standard, but without using lysozyme (RFC 59 standard points 6 and 7)
Data analysis with Cell Quest and Microsoft Excel [Cherry]
legend:
E.coli TOP10 constructs:
G- GFP
I- Invasin+GFP
J- J23102+Listeriolysin O+Invasin+GFP
A- pAraC+Listeriolysin O+Invasin+GFP
Hela cells with bacteria:
23-07-2010.007- without bacteria
23-07-2010.018- with G (1)
23-07-2010.019- with G (2)
23-07-2010.020- with G (3)
23-07-2010.015- with I (1)
23-07-2010.016- with I (2)
23-07-2010.017- with I (3)
23-07-2010.012- with J (1)
23-07-2010.013- with J (2)
23-07-2010.014- with J (3)
23-07-2010.008- with A (1)
23-07-2010.010- with A (2)
23-07-2010.011- with A (3)
cytometer plots:
FSC- Forward Scatter, cell size
SSC- Side Scatter, cell complexity
FL-1- FL-1 sensor, cell fluorescence
Data processed with Cell Quest:
Data from Cell Quest processed with Microsoft Excel:
legend:
Hela N- without bacteria
M1,M2,M3,M4- PIN Units (according to RFC 59 standard point 7)
SD- starndard deviation
ERR.-errror
more exact data:
