Week One:
6/13/05
Following OSHA training I returned to my lab to split an H460 or Lung Cancer cell line into additional flasks.
6/14/05
A Thin Layer Chromatography (TLC) assay was done and results obtained.
6/15/05
A plasma protien binding assay was done but was not optimal as only 13% of counts were freed following being centrifuged.
6/16/05
A plasma protein binding assay was again run and results obtained.
Week Two:
-June 20, 2005
An assay was done to evaluate the plasma protein binding of KPC-2. Results were unsuccessful as only an average of 13% of counts were recovered as nonbinding. The greatest amount of free counts is optimal as those that do not bind are available to circulate through the body system.
-June 21, 2005
The same assay was done as on June 20 to evaluate plasma protein binding ofKPC-2. Results again were unsuccesful as only 9.8% were freed from the serum spinnners.
-June 22, 2005
The plasma assay was repeated with caps being left off of the spinners in order to reduce capillary action, and in turn, free more counts. This method was not optimal, however, as only 12% of counts were freed.
-June 23, 2005
An assay was done to evaluate the plasma protein binding of KPC-2; however, adialysis bag method was used. This, in turn, resulted in some 86% of counts being freed.
Week Three:
-6/27/05
The protein plasma binding assay was repeated, but instead a dialysis bagprocedure was done. Spectrapor 2 dialysis tubing was used.
-6/28/05
The results from the 6/28 assay were obtained and was optimal as 100% of PBS counts were recovered and 86% of serum. An additional dialysis procedure wasset up and left overnight as it needs approximately 20 hours to equilabrate.
-6/29/05
Results were obtained from the 6/28 assay and was optimal as 100% of PBScounts were recovered and 83% of serum. Another plasma protein binding assaywas done comparing the equilabration of PBS and Serum at 37 degrees vs. PBSand Plasma at 4 degrees.
-6/30/05
In reviewing results from the 6/29 double comparison assay, serum was not affected by the changing of temperature from 4 to 37 degrees. Plasma assays,however, will be repeated as over 100%of counts were recovered, and is still in question.
Week Four:July 5, 2005
A plasma protein assay was set up. In order to do so new dialysis tubing was created. Following the start of the experiment, the plasma that was inserted into the dialysis bag was left to be spun overnight in order to evaluate equilibrium.
July 6, 2005
Results from the 7/5 plasma protein assay were obtained. They, however, were not optimal as all counts were not recovered; a similar problem like that found when using the centrifuge method. Another plasma assay was set up andleft overnight. Also HL60 cells were split.
July 7, 2005
Results from the 7/6 plasma protein assay were compiled. Another assay was not set up as it has to be read the next day. Values were better than those obtained on 7/5 and show that equilibrium can be evaluated correctly through the dialysis tubing method.
Week Five:6/11-6/14/05
The entirety of the week was spent compiling data. The results obtained were averaged and put into a summary chart which represents the average amount of drug inside and outside of the dialysis tubing which was the method used.
