Overall, the experience was positive. We all got a better understanding of how to micropipette because of the pre-lab reading. It was specific and provided enough detail so that we could move forward. In the beginning, the first and second stop was a little confusing. It was something that we needed to get hands-on experience with sot hat we could actually feel the first and second stop. While pipetting, we were fairly confident in our abilities after multiple trials. Even so, there was a couple of tubes that ended up with liquid still in there. Because of this, the actual volumes of those samples were slightly different. Our method for labeling was constant the entire duration.
Fluorimeter Procedure
Imaging set-up
We first got the Flourimeter box and opened the contents. We had to replace the phone stand because the original one was lower than what the experiment required. After, we got two plastic containers to prop up the device so that it was in line with the phone camera. Essentially, the phone needed to be held perpendicular to the slide and the camera needed to be parallel to the slide. Then we practiced taking a picture by putting a drop of H20 on the slide. We then closed the lid on the box and set the timer for the camera. While closing the side of the box, we covered any small cracks that light could get inside through to keep the inside dark.
Placing Samples onto the Fluorimeter
PLace the slides on the Flourimeter (Rough side up)
Add the pipette tip and set the volume to 80 microliters
Then push the plunger button to the first stop on the SYBR Green I tube
Submerge the tip in the liquid and release the plunger button
Transport the liquid from the tube to the slide (center lane, in between the two outer rows) by pushing down on the plunger button
Then push the plunger to the first stop on the 5 micrograms/ml tube of Calf Thymus DNA
Submerge the tip in the liquid and release the plunger button
Transport the DNA from the tube to the same spot as the SYBR Green I
Put the FLourimeter inside the box and turn on the light
Repeat steps 1-9 for the 2, 1, .5, and .25 concentrations
Data Collection and Analysis
Images of High, Low, and Zero Calf Thymus DNA
Calibrator Mean Values
Initial Concentration of 2X Calf Thymus DNA solution (micrograms/mL)
Final DNA concentration in SYBR Green I solution (µg/mL)
Patient 28137 : There was little to no images that displayed green flourescence
Conclusions
Patient 17649: Positive because the concentration of the PCR DNA was high which indicates a high fluorescence observed. The similarity of the Positive control further leads us to this Conclusion
Patient 28137: Slightly Positive because the concentration was barely positive but still lacked fluorescence observed.