BME100 s2015:Group2 12pmL6

BME 100 Spring 2015	Home People Lab Write-Up 1 | Lab Write-Up 2 | Lab Write-Up 3 Lab Write-Up 4 | Lab Write-Up 5 | Lab Write-Up 6 Course Logistics For Instructors Photos Wiki Editing Help
OUR COMPANY Name: Levi Riley Name: Alexandria Clark Name: Blossom Mendonca Name: Alina Kilic Name: Trevor Douglass Name: Andre Dang LAB 6 WRITE-UP Bayesian Statistics Overview of the Original Diagnosis System The division of labor consisted of 34 teams of 5 or 6 students who diagnosed a total of 68 patients. This allowed for the class to test a massive amount of patients in an efficient amount of time. Each group ran a PCR test on two patients, and prepared three samples of replicated DNA of each patient along with a primer mix to ensure accuracy. Each of the solutions were compared to both positive and negative controls. Each individual group ran its own positive and negative control when completing their individual PCR tests. The positive control would fluoresce if it contained DNA, while the negative control would do nothing because of its lack of DNA. If the sample did have DNA, it was compared to the positive control. If the sample did not contain DNA, it was compared to the H20 curve. To avoid error, a practice trial for using a micropipette was conducted so that it could be used with ease when testing with actual samples. This ensured the students accurately were able to pipette the correct amounts of mixes when using PCR so that the open PCR would work properly. Error prevention was also found in the fact that each group analyzed three samples from the same patient. This means there were 6 PCR tubes for only two patients (8 total when the positive and negative controls were included). This allowed for multiple trials to ensure that if one went awry, it wouldn't throw off all the data. After the PCR was complete, the fluorimeter was used in conjunction with SYBR Green I dye to test each sample of the PCR. In the fluorimetry, photos of the samples were taken and then analyzed through the use of ImageJ. ImageJ separated the photo into three different color channels (red, blue, and green), although only the green channel was used. The wavelengths of that green channel were compared to the light of the droplet in the test tube. That drop was analyzed to determine the density and area. There were three images per unique PCR sample, however, only the best picture of each sample was used for the ImageJ calculations. For ImageJ, an error prevention was put on in the fact that the Image J had to be calibrated based upon the positive and negative controls for each group. This means that each group is using its own standard which was created in their PCR machine. This ensures that the results will be accurate. Every group’s final data was then uploaded onto a document to compare the overall results and come to a conclusion. Overall, the class had a successful conclusion. On the other hand, this group’s conclusions were not successful as the data was inconsistent with the results. From the overall results, it can be deduced that the probability of having positive PCR results directly relates to having Coronary Heart Disease. What Bayes Statistics Imply about This Diagnostic Approach The results for calculations 1 where variable A meant there was a positive test conclusion while variable B showed the positive PCR reaction. Both individually showed the number of patients diagnosed and tested for both produced values that were less than .500 being relatively close to each other, indicating that Variable A and B correlates through the experiments. Now taking the Bayes values from both variables, A for positive test conclusion and variable B for positive PCR reaction where the probability of variable B given A shows that the Bayes value is close to 1.00 or 90%, which means that majority of the patients that did test positive to the PCR reactions did in fact also test positive for cancer. Switching the variables where the probability of Variable A given B where the patients that concluded positive for coronary heart disease given having positive PCR reactions equaled 94%, a higher frequency then the probability of B given A. This concludes that the number of patients that tested positive for either PCR reaction or Positive Coronary heart disease has a high probability of testing positive for the other. The results for calculation 2 where Variable A meant there was a negative test conclusion while variable B showed the negative PCR reaction. Both variable A and B showed the number of patients diagnosed and tested negative produced Bayes values that were slightly above .500 being 2% different from each other, indicating both negative test results correlate with each other through the experiment. Taking the frequency from both variables, where the probability of having a negative PCR reaction given negative test conclusions shows that both variables correlate with each other having a probability of 94%. Inversely, the probability of a negative PCR result given the conclusion being negative gives a probability of 90%. In conclusion, a negative Final Test Conclusion and Negative PCR reaction does in fact correlate with each other. The results for calculation 3 where Variable A meant the total patient that will develop disease while Variable B shows the positive test conclusion. Variable A producing value of 37% and variable B producing a value of 43%. Now the probability of variable B given A, Frequency of positive test conclusion given patient that will develop disease, gives a value of .9 or 90% means that the number of patients that do test positive will have a high probability of developing the disease. Now the probability of variable A given B, total patients that will develop the disease given the probability of positive final test conclusions, gives a value of .77 or 77% means that the patient that will develop the disease later on will test positive during the final test but not completely conclusive. Since the value is not close to 1.00 there were errors in the value or the actual experiment. The results for calculation 4 where Variable A meant the total patients that will not develop the disease while variable B shows the total negative test conclusions. Variable A showed the Bayes value being close to .500 or 56% and Variable B shows a value above .60 or 62%. Taking the frequency for both values where the probability of variable B given A, negative Test Conclusion given Total Patient that will not develop disease, gives a Bayes value close to 1.00 or 94% which shows that yes, patients that test negative will be likely to not develop the disease. Now the probability of variable A given B, total patients that will not develop the disease given the probability of the negative final test conclusion, gives a frequency that is 84%. Meaning that majority of the total patients that do not develop the disease will be more likely to have a negative final test conclusion though not completely conclusive. Computer-Aided Design TinkerCAD The tinkerCAD tool is an online interface to create 3D objects and devices. This interface can be used to create simple objects or to create intricate designs. During this lab, TinkerCAD was used to create part of the open PCR device container(front-back-bottom and one side). The rest of the PCR machine would be assembled in a likewise fashion; however, the lab TA told the group to only assemble 3 sides for this portion. The TinkerCAD for the basics of the PCR Machine is as follows: Our Design The new design was chosen to address the problem regarding amount of samples which can be tested. The original openPCR device only could fit 16 micro-tubes, which only allowed for 16 tests, and in the application of this course, only allowed for two teams per box. This new design will expand the amount of tests per box, allowing for more tests, the new openPCR design has double the area for microtubes, allowing for double the amount of tubes. The new design will accommodate up to 32 micro-tubes. In order to do this, the group assembled the outside of the Open PCR machine and increased the length and width so that the dimension created a doubling of the inside area. This doubling in area means that double the amount of test tubes (and double the amount of samples) will be able to be tested. Feature 1: Consumables Kit The consumables that will be packaged in the kit will be: PCR reagent mixers test tubes to hold samples 32 test tubes which are 50 μL of content in each: Mix contains Taq DNA polymerase, MgCl2, and dNTP’s DNA/ primer mix, 32 tubes, 50 μL each: All tubes contain the correct reverse and forward primer 32 empty PCR tubes Pipette tips Micropipettor SYBR Green I Dye Glass Slides For Fluorimeter For the way in which these consumables will be packaged, they will come in a box which contains each section divided appropriately so that each consumable will be grouped together. The box will have a flip-top lid which moves backwards and has printed labels at the top which identify exactly which consumable is in each compartment. The consumables will be grouped together and allow for the production of thirty two PCR reactions. This means that the contents will be enough to fill the newly designed PCR machine for one full run in which each compartment is entirely filled with an individual PCR tube. The benefit of this is that the new consumables are organized and easy to keep track of that so that the experimenter will never get the components mixed up. The new feature that this group is offering is also the ability to order specific primers in the consumables package so that the PCR can be ran for whatever the consumer needs. Also, the new pipette tips will allow for reuse so that they do not need to be thrown away. When consumers order new consumable packages, there will be an option on whether to include or not include pipette tips. This is a customizable feature to help the consumer and allow them to spend less money in the long run. A major problem with the consumables in our kit used in this experiment were the amount of plastic disposed. As engineers, sustainability should always be a top priority when conducting an experiment. As a solution to this problem, the plastic tubes used for micro-pipetting should be produced using more durable material and eco-friendly materials. This material will have to be one where it will not absorb the sample or liquid that it is picking up, so as to not contaminate later samples. With more durable and eco-friendly materials, these plastic tubes can then be sanitized and reused multiple times before disposal, thus lessening the amounts of plastic waste. Also, with eco-friendly material, the decomposition of these tubes will not contribute to rapid increase of plastic pollution in the environment. Feature 2: Hardware - PCR Machine & Fluorimeter The PCR machine is used to replicate the DNA multiple times, the machine in this system can be redesigned to address multiple problems. The PCR machine only allows for 16 micro-tubes to be replicated during each use, each use also takes 2 hours to process through and must be connected to a computer the entire time. These requirements waste time for the user, and are inconvenient. The PCR machine can be redesigned to better adapt to the users needs by allowing for more samples in each run, and by making each of those runs faster and not needing a computer connection. If the PCR machine was to be enlarged, it would allow for more samples to be included in one 2 hour run. This change will allow for the user to use less time to replicate a certain quantity of samples, and allow for less PCR machines to be needed for that quantity of samples. Making the replication time faster could be done by increasing the components which cool the samples, so it takes less time to cool. Even if the PCR machine continues to take 2 hours to run, it would be more convenient to the user if it did not need to be hooked up to a computer, for analysis to be sent. The PCR machines requirements to be connected to a computer the entire 2 hours, means the user cannot use their computer during the run, which is inconvenient for them. It would be beneficial if the PCR machine had an internal storage device for all the information to be stored, and later transferred to a computer using a USB device. All of these changes would allow for a smoother run, which has the least negative impact on the user. The flourimeter is used to analyse the DNA after it has been replicated in the PCR machine. This processes is long and requires carefully taking a picture, then quickly shutting the door to a dark cube where the DNA is located, this picture then must be loaded unto a computer with imageJ and analyzed. The flourimeter system has many issues, the box could not be shut soon enough, the phone has to be stacked so it is level with the DNA drop, the picture could be blurry, and it takes time to analyze each photo on imageJ separately. If the flourimeter was incorporated into the PCR machine, alot of setup time and human error would be eliminated. If under the PCR machine, a fourimeter system was set up, and the pictures automatically taken, there would be no need to worry about shutting the door or human error which could arrise. This could be done if, after each sample finished replicated, they dripped unto a slide where a camera in the PCR machine would take a picture of each sample, and sent it to a storage bank which can be accessed later. Another option is to create an internal picture device, so a button could be pushed on the outside, which would take the picture, eliminating the risk of outside light effecting the picture. To fix the time and effort needed to analyze each picture, a separate program could be created to upload each picture, split the color channels, and can recognize where the droplet is on the picture, from there it would be able to analyze the pictures for area, density, and any other information needed, then produces a large list of this data for all the pictures uploaded. These changes would allow for the fourimeter analyses process to happen faster and without as high of a change for error. The new device designed looked at the PCR machine and addressed the issues of quantity of samples analyzed at once. The current design only accommodates 16 micro-tube samples at a single time. This means that the user had to either run the machine multiple times or buy multiple machines if they wanted to be able to analyze more than 16 samples. The new design is clean and cost effective, by simply enlarging the PCR machine, a larger base for samples is allowed. This base will be able to accommodate more samples, and allow for a faster samples/hour rate of replication. The design created will have twice as big of dimensions as the original PCR machine. Being twice as big, it will allow for twice as many samples, meaning 32 samples will be able to be accommodated for replication at a single time. This new designs significance is that it allows for the user to replicate more DNA in a single run, so that with one machine they can produce what two machines, or two runs used to, now the user will only have to do one run for 36 samples, and will have to do less total runs, or use less total machines.