Safety

Safe Project Design

Living Organisms

Our project did not include any hazardous organisms. The live material that was used is all on the iGEM white list and are widely used in synthetic biology research by our lab.

• Bacteria: Top10, XL1Blue; two E.coli variants.
• Mammalian cell lines: HEK293T/17

Parts

The new part that our team introduced is the coding sequence for FVIII. This protein is not dangerous to human, animal or environmental health. It could only be dangerous if the protein is administered to patients at a too high dose, which would lead to thrombosis.

Future applications

If our project were to be developed further into a usable therapeutic, it would be able to help patients with hemophilia. It would improve their life with a less invasive therapy. With further development, certain safety aspects should be addressed. First of all, even though they are considered safe, the toxicity of the nanoparticle components needs to be thoroughly assessed before they can be used in any human trials. In addition, the dose of the medication should be carefully considered, because too high doses of FVIII could increase the chance of thrombosis. The engineered organisms are a part of the manufacturing process, but would never be released beyond containment so no special considerations have to be made there.

Chemicals

Due to the nature of our project, hazardous chemicals were used for multiple applications. However, the final product does not contain any of these chemicals.

• Dichloromethane/methanol: used for purifying the synthesized ionizable lipids.
• Phenol/chloroform: used for RNA purification.
• Ethidium bromide: used in gel preparation for electrophoresis

These chemicals were used in a fume hood and gloves were worn when necessary.

Lab Safety

Biosafety and chemical safety

When working with biological materials and possible infectious agents, it is important to keep track of proper safety etiquette. There are 4 levels of biosafety (BSL) and chemical safety (CSL) currently used in the research fields with level four being the most hazardous. Our project consisted of experiments conducted in BSL-1,2 and CSL-1,2 level.

Our research facilities

All facilities for our project were provided by Radboud University.

Chemistry and biology labs

All the lipid nanoparticle experiments were carried out in the laboratories of the Education Institute for Molecular Sciences (EIMS) in the Faculty of Science, Radboud University. In these laboratories, it is required to wear 100% cotton lab coat and safety goggles at all times when present in the labs. Additionally, these lab coats must be taken off when exiting the rooms.[1]

The laboratories are equipped with multiple fire safety measurements. In the designated areas these items can be found:

• carbon dioxide fire extinguishers;
• foam fire extinguishers;
• buckets of sand;
• fire blankets.

Each of these are used for extinguishing specific types of fires, for example, sand for fires consisting of metal reagents. In addition to these fire safety measures, multiple red panic buttons can be found which, upon activation, result in electricity shutdown in the entire lab area. In the events of a large hazardous chemical spill on the skin or in the eyes, multiple emergency showers and eyewash stations are available throughout the lab for immediate use.[1] When working with mechanical equipment, such as desiccator, prior explanation was given by our instructors to ensure proper usage and safety towards the equipment.

Hansen lab

Part of our research process was carried out in the Hansen lab of the Biophysical Chemistry Department in the Institute for Molecules and Materials, Radboud University[1].

All safety equipment mentioned in the previous section is available in this lab. Our members who carried out experiments there, followed the measures of proper lab safety. Only difference, it was not a requirement to always wear goggles. Additionally, most experiments were carried out in the presence of our instructor Frank Nelissen.

Chemical Waste

All waste bins with liquids are equipped with funnels to reduce chemical evaporation. All waste bins can be found in fume hoods. Except, in biochemical lab, the large solid waste bin can be stored outside of fume hood. All uncontaminated solid waste can be thrown in multiple normal waste bins found in lab.

Personal safety

Lab coats were worn closed at all times during experiments. All participants wore clothing that covers ankles and is not made of highly synthetic materials. Also, those with long hair, had their hair tied up. All footwear worn was closed, not open. When working with more hazardous and carcinogenic substances, plastic disposable gloves were worn until the experimental had been carried out. After, the gloves were disposed into chemical solid waste. The lipid nanoparticle research was always carried out in the fume hood due to the hazardous levels of the substances. These fume hoods were always kept closed if not in use. All students who carried out research in these labs were introduced to the chemical safety equipment, measurements and taken a safety exam provided by their university. A part of the team has also had official biosafety training as a part of their university training. The rest of the team received the same instructions regarding biosafety.

References

1. Hansen lab. (n.d.). Hansen Lab. Retrieved from https://www.thehansenlab.com/hansen-lab-radboud-biophysics-team
2. Code of Conduct for Lab Courses. Radboud University; September 2022.