BME100 f2014:Group23 L6
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Lab Write-Up 1 | Lab Write-Up 2 | Lab Write-Up 3
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LAB 6 WRITE-UP
Overview of the Original Diagnosis System
For testing the 68 patients each of the 34 teams of 6 students tested 2 patients. For each patient 3 DNA samples were taken to avoid any coincidental errors. Each team also had positive and negative controls, as well as different concentrations of the DNA sample in order to be able to analyze whether each patient sample was positive or negative. Three images of each concentration and each patient sample and positive and negative control samples were taken, so as to prevent error. In order to set up the fluorimeter a camera was leveled to each sample drop combined with SYBR green. These images were analyze for numerical values of green INTDEN using ImageJ, since green fluorescence determines the prescence of the disease. These numerical values allowd us to analyze the concentrations of the patient samples and compare them to the controls. If all 3 or 2/3 samples for each patient where determined to be positive (that means within the positive concentration range) then the teams concluded a positive result for that patient, and so with negative results. After analyzing each image, 8 patients received an inconclusive test result, while 30 patients received a positive test result, and 24 patients received a negative test result. Additionally, 3 groups did not complete the test analysis, so 6 patients did not receive any results.
Calculations 1 and 2 show that the system is not perfect, but still reliable for testing this disease. Both specificity and sensitivity calculations were close to 100%, and were at worst 23% from being perfectly accurate. As for human error there could be errors in concentrations in the drops for the fluorimeter. Another possible human error could be cross-contamination by using the same tips while transferring different samples. Another possible error, both human and machine, could be faulty INTDEN values from the ImageJ software. This could be due to wrong placement of ovals for the measurements.
Name of device: PCRF- Polymerase Chain Reaction Fluorimeter
The new design is quite simple, it combines both the fluorimeter and the PCR machine into one, and the fluorimeter is improved for easy access and to make the overall process faster. The design includes an added structure on the side of the PCR machine, a box containing four structured layers. It is essentially the original fluorimeter except compressed onto four levels. Each level can be opened, and a slide with a drop can be inserted. On each end is an opening for the camera. The side can be lowered, immersing the four layers in darkness, and the camera can be moved up to each of the four openings, allowing four drops to be photographed simultaneously. In addition, a camera holder can be inserted into several locations along the wall to hold the camera steady for the photograph of the drop. The camera will be provided, along with a cord connecting it to the computer. There will also be a software program that allows for instant calculation of the necessary values for the fluorimeter process. It would be relatively cheap to include with the original PCR machine, especially since the entire design can be taken off and packaged separately. It is simple, and allows for the entire process to run smoothly and quicker than the original fluorimeter design. We used the TinkerCAD to design this extra portion that could be added onto the PCR machine, and the four layer contained inside the design.
Feature 1: Consumables Kit
All the materials will be placed in a dark cold box. All components will be wrapped in bubble wrap as a precaution. The bubble wrap will make the product easy to store and make it safe from any situations when it is transferred from area to area. The bubble wrap will be wrapped around the PCR machine twice in order to provide ultimate protection. The dark cold box protects the positive and negative samples as well as the SYBE GREEN 1 Solution from any spillage. The flourimeter slides and pipette tips will back packaged with the PCR machine and all the contents it contains but in a separate box in order for it to be more protected. It is important to consider that the flourimeter slides are glass and are delicate to any kind of movement. The box also protects the products so they stay sanitized and away from any type of dirt that it could possibly contact.
It is important to make sure a PCR machine and whatever contents it contains are safe from any type of damage since these are delicate products. Which is why precise packaging is necessary in order for the product to transition from one are to the next. This is why detailed packaging is important due to the fact that specific substances as well as other delicate products such as the pipette tips and flourimeter slides should be packaged with precaution.
Feature 2: Hardware - PCR Machine & Fluorimeter
The design of the PCR machine in terms of materials, with the wood exterior being durable, lightweight, and fairly resistant to any heat produced, was kept the same. Likewise, the overall structure of the PCR was kept the same. Instead of redesigning the PCR machine, the flourimeter was attached to the side of the PCR machine. The flourimeter consists of a rectangular box with four levels. The side of the box can open so that four slides may be placed inside, one on each level. A camera will come attached by an adjustable camera holder that may slide up and down to take pictures of the droplets. This design is ideal because all of the work can be done in one space. Additionally, a software to analyze the images will be included in the package. This way, by hooking up the camera to the computer, the software will automatically analyze the images and come up with a positive or negative result, so long as each picture is labeled correctly. To ensure this, the device will ask that the specific drops, four at a time, are all placed on the correct level. For instance, the device would first ask that the positive control, negative control, patient 1 sample 1, and patient 1 sample 2, with SYBR Green mixed in each droplet, be placed on each level. The samples should be placed on each level in the order that they are asked for, with the positive control on level 1, the negative control on level 2, and so on.
Once all droplets are placed correctly, the person using the device would press the start button for the flourimeter portion of the device. Then the camera would automatically take three pictures of each level, starting at the bottom so that each picture corresponds to the correct sample, and move upwards until all samples have been accounted for. Then the next four samples would be asked to be placed on each level (patient 1 sample 3, patient 2 sample 1, patient 2 sample 2, and patient 2 sample 3), and the camera would begin taking pictures again when the start button is pressed again. This process would be repeated until all samples are accounted for. Then the camera would be hooked up to the computer, and the pictures would be transmitted. The software will then analyze the SYBR Green in each droplet to make a positive or negative conclusion. The software would already have the analysis values for the different concentrations of the fluorescence, so the person using the device would not have to take pictures of the different concentrations.