Proposed Implementation

Who are your proposed end users?

DIAS diagnostic IVD test can be used by physicians as a part of the annual blood-based check-up examinations in a portion of the population showing high risk for NSCLC such as current or former heavy smokers at age 50 to 80 or patients with verified elevated risk for lung neoplasms. Furthermore, DIAS can be used in any other case of suspicion for lung cancer after clinical examination and doctor's instructions. Who could be the end users of the DIAS diagnostic device? Definitely the recommended users of our diagnostic test could be the NSCLC-risk groups and other patients referred by their doctor with lung cancer suspicion.

How do you envision others using your project?

As scientists dedicated to cancer research, our envision is to detect NSCLC cancer development at risk groups even at the early stages of the disease. Doctors could encourage risk groups and patients with suspected lung cancer to do the DIAS IVD test annually and at the time required in a proper facility such as a hospital or a diagnostic center. The result of the test will reveal a lung cancer possibility score. Afterwards, physicians will evaluate the displayed score and decide whether the patients should be referred for further more invasive tests.

How would you implement your project in the real world?

For the practical implementation of the DIAS diagnostic IVD test in real world situations, our system's handling method is separated into two main procedures. The first procedure requires the premixing of the appropriately pretreated blood sample with the DIAS detection master mix which contains all the necessary reagents in specified preformulated proportions (figure 1). More information regarding the DIAS detection master mix can be found on the measurement page.

The following procedure requires the test sample entering into a droplet centrifugal microfluidic cartridge which is incorporated in a standard falcon tube, shown in Figure 2 . After processing at a fixed centrifugal acceleration at a standard laboratory centrifuge, an emulsion is generated with fluorescent water-in-oil droplets. Afterwards, the cartridge is removed from the falcon tube, and the emerged emulsion is ready to be transferred in the Neubauer improved counting chamber for droplet read out. Fluorescence images of the generated droplets can be easily obtained by a simple fluorescent microscope and automatically analyzed by a visualization-quantification software. Based on the software analysis, the number of green fluorescent droplets could be easily correlated with the amount of the target miRNA presented in the patient's blood sample. The amount of the target miRNA in the sample will finally reveal the lung cancer probability score of the patient who conducted the DIAS examination.

What are the safety aspects you would need to consider?

Considering the safety aspect of this project, the diagnostic procedure does not endanger the patient since it only requires a blood sample. This examination can take place only in appropriate centers such as a hospital or a diagnostic center. Thus, the patients' examination will be conducted by qualified staff for their thorough monitoring in appropriate conditions. Also, the visualization-quantification software will automatically provide the lung cancer probability score preventing errors in the elucidation of the results. Although, concerns could be aroused by patients in case the IVD test fails displaying a false positive or false negative result. For this reason, we communicated with experienced people in the field by contacting the International Organization of Medicines of Greece. More information regarding this edifying discussion with the competent regulatory authorities are provided in human practices page.

What other challenges would you need to consider?

Two of the fundamental challenges in IVD tests constitute the cost and the conduct procedure of the test. Due to the construction of the microfluidic chip through the 3D printer the manufacturing cost and finally the cost that patients need to pay for the test is significantly reduced. Regarding the test handling procedure, the centrifugal microfluidic cartridge requires the usage of centrifuge equipment available in all diagnostic centers and hospitals. Even though we had no time available to test our project with this structure, we managed to simulate the flow system of the centrifuge utilizing a syringe pump system. The syringe pump system enables a controlled flow rate of the samples. Through the microfluidic device, the test sample (aqua phase) will be mixed with an oil phase and due to the architecture of the microfluidic chip, the final sample will be shaped as a water in oil emulsion. The final sample will then be loaded onto a Neubauer improved counting chamber to count the fluorescent droplets. The droplet generation system with the syringe pump is displayed in Figure 3.