Monday 6-14:  Make a blanked data sheet for data collected today and up till yesterday for experiment 3.Day 4 collected data is consistent with data presented before today.  We see a significant increase in cell absorbance after 60 minutes.  Blanked data average absorbance comes to .8542 and .2808!!!  These results are on the order of previous experiments conducted by Go-Shau but how long will the media last??
    I am also trying to standardize the MTS incubation period.  A 3 and 4 hour MTS incubation idd not show statistically different results.  Therefore, 1 hour is the best time period.  I am still analyzing 24 hour results and they show some promise because they yield greater consistancy between replicate wells.  A slight statistical anomaly has occurred because some values are extreme.  I am looking for statistical measure to preserve these values. (T-Test??)

Tuesday 6-15-99: MTS reading looks great.  Huge difference (>200%) seen between treatments.

Wednesday 6-16:  Reading still looks good.  Started planning next experiment which will involve C.neo inhibition and killing by Macs induced with GMCSF!

Thursday 6-17-99: Both numbers went up as I read the Day 6 plate for experiment 3.  This does not correspond to previous results.  Reasons why this could occur:
        1. We use DMEM instead of RPMI. DMEM has higher glucose concentrations.
        2. We used 30,000 cells per well, instead of 50,000.  Higher values should be expected.

Friday 6-18-99: Last day of experiment 3.  Day 7 reading indicates both went up again.  But GMCSF treated media is definately depleted.  THE RESULTS FOR THIS EXPERIMENT ARE SHOWN BELOW:
        Note:  The data is an average of all wells and is statistically adjusted to eliminate errors of grater that 50 percent.  No such values were excluded!!

 

I also started a new experiment today, hereafter referred to as experiment 4.  I did another macrophage extraction from a 1 month old rat.  I will test the C. Neo killing ability of Macs treated with or without GMCSF.  The colony plating procedures will be taught on Wednesday by Go-Shau.  2 time points for the C.Neo infection will be used (24 and 48 hours)  My mac extraction went as planned with a small yield (small rat) of 3.5 million cells.  The plate plan I used is shown below.  It will be be incubated for 5 days.

The number in the well represents the concentration of GMCSF (ng/ml) used.  83,000 cells/well were plated.  One row was skipped between rows of cells.

                24 hour time point                                   48 hour time point