Safety

"Whenever I'm about to do something, I think, 'Would an idiot do that?' And if they would, I do not do that thing." Dwight, The Office

GENERAL LABORATORY SAFETY


Safety is an important factor when talking about scientific research. It is necessary to minimize the risks and costs that can cause laboratory-related dangers, for both the researchers and the results of the experiments.

In order to ensure laboratory safety, all the members of the team underwent specific training concerning security and safety measures in the laboratory. Some of us had already worked in molecular biology laboratories and had a solid background in safety measures. Moreover, all the members of the laboratory were provided with personal protective equipment, such as gloves and different laboratory coats, one for the general laboratory and the other one for the room containing the biosafety cabinets. The laboratory coats used while working in the biosafety cabinets never left the room and our clothing was disinfected and sterilized to prevent contamination of the external environment.

Our workspace was classified as Biosafety Level 2 (BSL-2), covering laboratories that work with agents associated with human diseases and that imply a moderate health hazard. The practices required in a BSL-2 laboratory setting and that our team followed are listed below.

  • Mechanical pipetting only (no mouth pipetting allowed).
  • Safe sharps handling.
  • Avoidance of splashes or aerosols.
  • Daily decontamination of all work surfaces when the work is completed.
  • Hand washing.
  • Prohibition of food, drinks, and smoking materials in the laboratory setting.
  • Personal protective equipment, such as: eye protection, gloves and a laboratory coat or gown.
  • Appropriate personal protective equipment (PPE) must be worn, including laboratory coats and gloves. Eye protection and face shields can also be worn, as needed.
  • All procedures that can cause infection from aerosols or splashes are performed within a biological safety cabinet (BSC).
  • An autoclave or an alternative method of decontamination is available for proper disposal.
  • The laboratory has self-closing, lockable doors.
  • A sink and eyewash station should be readily available.
  • Biohazard warning signs.

Immediate decontamination after spills was required and used materials were decontaminated through an autoclave. All the used materials were properly trashed into each specific container to ensure optimal waste management.




SAFE PROJECT DISIGN


Our team worked with E.coli DH5-alpha to amplify and select plasmids, which afterward were transfected to HEK293T cells. The main purpose of their use was to generate a stable cell line containing features that improve the production of exosomes. E.coli DH5-alpha is not considered hazardous by the 2012 OSHA Hazard Communication Standard (29 CFR 1910.1200), as this reagent does not contain substances considered to be health-damaging at their given concentration. Conversely, the HEK293T cell line requires working more cautiously, although it is only able to proliferate under specific laboratory conditions and cannot spread autonomously in the environment, infecting humans or living organisms.

Taking into account the therapeutical intentionality of our exosomes, the project would need extra support to manage the identified risks before accessing the commercial market, with special relevance in the regulatory and clinical trials' design fields. Moreover, our kit could only be used in laboratories that have the resources to follow all the safety measures and steps that might be needed.




SPECIFIC LABORATORY SAFETY


Our team worked under a biosafety cabinet, which purifies the air of potentially infectious aerosols and animal dander. Each member of the laboratory team read and signed a self-responsibility document provided by the laboratory of the Josep Carreras Leukaemia Research Institute to be aware of the proper use of this type of cabinet. Some of the experimental techniques that we performed also required the use of UV light or other health-hazardous chemicals; thus, additional safety protocols were needed:

  • UV light: UV light was used to sterilize the laminar flow hoods and during the DNA purification from agarose gels. The possible hazards of encountering UV light were minimized with the correct use of the equipment. As an example, the cabin was always properly closed to ensure its retention.
  • Antibiotics: kanamycin was used during the cell-selection procedure. According to their material safety data sheet (MSDS), ampicillin, chloramphenicol, and kanamycin are reported to have hazardous effects at high concentrations and both acute and chronic exposure to humans via inhalation, ingestion, or contact with the eyes. The possible hazards of working with kanamycin were minimized with the correct use of personal protective equipment when handling the compound.
  • Carcinogens: methanol, SYBR-Saf, and Trypan blue, which are considered to be carcinogenic via direct contact, were used during our project. The possible hazards of working with them were minimized with the correct use of personal protective equipment (e.g. gloves) when handling the compounds.

Our team followed the safety and security legislations of our country, at the Spanish and European levels. Laws and regulations that must be followed in our region can be found in the following links:

ARCHIVO Y PRINCIPIOS DE BUENAS PRACTICAS DE LABORATORIO
OECD SERIES ON PRINCIPLES OF GOOD LABORATORY PRACTICE AND COMPLIANCE MONITORING