We used the pipettor to transfer the PCR reaction mix, patients 1 and 2's replicate mix, the positive DNa/primer mix, and the negative control. The key to this was proper technique. When drawing fluids, only push the pipettor to the first stop, but when ejecting fluids, push all the way to the second stop. Failure to do either of these will result in a difference in the actual amount as compared to the desired amount of substance. Next, we placed the tubes with the substances into the OpenPCR device.
Fluorimeter Procedure
Smart Phone Camera Settings
Type of Smartphone: iPhone 6S
Flash: Off
ISO setting: Auto
White Balance: Auto
Exposure: Auto
Saturation: Auto
Contrast: Auto
Camera set-up
Distance between the smart phone cradle and drop =
Placing Samples onto the Fluorimeter
Set volume on pipettor to 80 microliters.
Place slide into fluorimeter such that the holes that will be used are aligned with the blue LED light.
Firmly attach tip to pipettor.
Pipette 80 microliters of SYBR Green I onto one of the middle holes of the slide.
Eject pipette tip into waste basket.
Firmly attach another tip.
Pipette 80 microliters of .25 concentration of the calf thymus into the hole right above the SYBR Green I.
Use timer on camera to take pictures of the solution in the dark under the blue LED light.
Dispose of the liquid on the slide into the liquid waste basket.
Repeat steps for the .5, 1, 2, and 5 concentrations of the calf thymus.
Data Collection and Analysis
Images of High, Low, and Zero Calf Thymus DNA
Calibrator Mean Values
Calibration curves
Images of Our PCR Negative and Positive Controls
PCR Results: PCR concentrations solved
PCR Results: Summary
Our positive control PCR result was 3986962.3 μg/mL
Our negative control PCR result was 2182675.3 μg/mL
Observed results
Patient 78778:
Patient 63691:
Conclusions
Patient 78778: positive because values above positive control
Patient 63691: positive because values above positive control
BME 100 Fall 2015
