Introduction

With SCENTIPD, we are dedicated to advancing the field of Parkinson's Disease diagnosis. Our objective is to create an innovative, practical, and cost-effective diagnostic kit. To achieve this, we employ a unique sensor technology centered on bacteriophages, designed to detect specific Volatile Organic Compounds (VOCs) present in a patient's skin sebum.

Throughout our scientific journey, we have successfully achieved three key milestones, each characterized by innovative modifications:

Substrate Development: Our first milestone involves the development of specialized substrates. These substrates provide the foundation on which our genetically engineered phages create distinctive structures, resulting in observable color changes. This process represents the initial crucial step.

Electronic Pulling Device: In our second milestone, we innovatively repurposed a syringe pump to achieve precise manipulation of the substrates and facilitate the interaction between the phages and VOCs. A pair of tweezers was vertically mounted on the device to securely hold the delicate phage wafers, enabling the fine control necessary for our diagnostic kit.

Integration of 3D Printing: A significant juncture in our journey was the successful integration of 3D printing technology. This enabled us to design and produce protective enclosures for our substrates. These enclosures serve a dual purpose, providing protection for the substrates and ensuring their seamless operation and compatibility within the diagnostic kit.

Fabrication of Gold-Coated Silica Plates

In our pursuit to develop an innovative diagnostic kit for Parkinson's Disease, we recognized the need for specialized gold-coated silica plates. These plates would serve as crucial substrates for our sensor technology, enabling the detection of specific Volatile Organic Compounds (VOCs) in a patient's skin sebum. This fabrication process was carried out under the guidance and expertise of Dr. Eleni Efthimiadou and her laboratory in the field of Inorganic Chemistry at the Department of Chemistry, National and Kapodistrian University of Athens.

The fabrication was facilitated according to “Gold coated silica plate fabrication protocol”. Since it was the first time we followed the procedure every step was documented as shown below:

Note: The following protocol outlines the meticulous fabrication process that resulted in the creation of 15 gold-coated silica plates.

Repurposing of a syringe pump as an electronic pulling device

A pivotal phase in our experiment centers on the meticulous self-assembly process of phage-based sensors. This step demands a precise lowering and raising of gold-coated slides from a phage solution at controlled, gradual speeds. The purpose of this precision is to leverage the meniscus of the solution to facilitate the formation of desired nanostructures on our fabricated substrates.

Initially, we sought a programmable syringe pump, similar to the one employed in the protocol we followed. Unfortunately, our laboratory lacked this equipment, prompting the need to fabricate a specialized pulling device for precise substrate manipulation. We initiated the design process by outlining the device's requirements. It became evident that a simple one-axis structure, capable of moving a gantry along a lead screw with precision, met our needs. To estimate the cost of fabrication, we compiled a rough bill of materials. A significant challenge arose concerning the required pulling speed and motor torque, with the process demanding speeds between 10 and 300 µm/min. After comprehensive research, we determined that a high-torque stepper motor, coupled with a microstepping-ready motor driver, would yield the desired results.

However, instead of proceeding with our initial plan, we stumbled upon a more efficient solution that circumvented the costs and time associated with creating a DIY pulling device. A generous contribution from a lab at the Hellenic Pasteur Institute provided us with an NE-1000 One-Channel Programmable Syringe Pump. This device met all our requirements, offering precise movement along a single horizontal axis at sustained low speeds, and it was programmable to suit our needs.

This programmable syringe pump had the capability to be programmed through a computer using specialized software and RS232 protocol communication or directly via the pump's interface. Initially, the direct computer interface seemed straightforward, allowing us to create the necessary phases for the pulling program and load them onto the pump. However, communication between the pump and the computer presented challenges. The RS232 standard for serial communication and data transmission necessitated a specialized cable. Several cable combinations, such as USB to DB9 and DB9 to RJ11, were suitable for our specific use. Yet, we determined that the most efficient approach would be to acquire a direct USB to RJ11 cable with an FT232 chip to convert the signal, adhering to the RS232 standard. After procurement, we modified the cable as the wiring on the RJ11 connector did not match the pump's port pinout. We adjusted the core arrangement by cutting and crimping a new RJ11 connector at the cable's end. Despite installing all required drivers, we encountered difficulties establishing a connection with the pump. While the computer recognized the new virtual RS232 connection, the pump remained unresponsive to our commands. Consequently, due to time constraints, we opted to program the device directly through its interface.

To attain the desired results, we calculated the flow rates the pump would need to operate at since it couldn't accept standard speed rates as input. After selecting a syringe an inner diameter of 14 mm, we developed the following program:

The pump was positioned vertically, enabling the pushing block to move and withdraw the substrate from the phage solution. To secure the wafer, we utilized a pair of stainless steel tweezers, two binder clips, and a rubber band, providing a stable foundation for the lightweight substrate.

Completing the hardware ensemble for our diagnostic kit involved the design and 3D printing of a specialized enclosure using. This enclosure was thoughtfully crafted to accommodate five of our carefully prepared self-assembled phage sensors.

Design and Dimensions: The enclosure takes the form of a compact box with an open top, designed to provide easy access while ensuring the security of the sensors within. To facilitate visual assessment of color changes, the enclosure features a closing mechanism that involves sliding a suitable piece of acrylic. This design ensures that the transformative color shifts in the sensors can be readily observed and assessed.

The inner dimensions of the enclosure were optimized at 3 x 5.5 x 2 cm, aligning precisely with the saturation requirements for the enclosure with the relevant Volatile Organic Compounds (VOCs) in our diagnostic process. The entire design process was executed using Tinkercad, a user-friendly 3D design platform, ensuring both functionality and ease of use.

Printing and Material Selection: The enclosure was brought to life through FDM 3D printing technology, utilizing PLA (Polylactic Acid) as the chosen printing material. PLA was selected due to its cost-effectiveness and ease of printing, making it highly suitable for the rapid prototyping phase of our project.

Accessibility and Collaboration: For those interested in utilizing our enclosure design, the STL file can be conveniently accessed through this link