Knowledge and awareness of rules for personal and biological safety are highly important to creating a safe, organized, collaborative and productive lab environment.
Working in the laboratory requires constant surveillance for hazardous materials and practices. It also requires that we be able to identify potential hazards and prevent or manage them properly without compromising our safety and the safety of others. Lab etiquette informs us on how to behave properly in the lab by developing safe practices and habits, as well as effective responses to potentially hazardous activities and outcomes.
Disposal and Spills
- Place used or contaminated lab supplies in proper waste containers. Locate the regular trash, biological waste containers, sharps and broken-glass containers. Any lab supplies that come into contact with cultures, media, reagents or any potentially hazardous material should not be left on the benchtop.
- Place used reusable equipment in proper receptacles for autoclaving; this includes metal loops, hockey sticks, glass beads, forceps, and glassware.
- Dispose of samples, cultures, and media in designated biological waste receptacle.
- Dispose of used toothpicks, wooden sticks, plastic tips, weigh boats, Petri dishes, and other disposables in designated biological waste receptacles.
- Dispose of used sharps (slides, coverslips, Pasteur pipettes, broken glass, etc.) in designated sharps and broken glass container.
- Ask your instructor about glass (e.g., microscope slide) and/or broken glass disposal.
- Properly dispose of contaminated media and plates.
- Do not remove the lid off plates with fungal growth. Plates with yeasts or molds disseminate spores, which rapidly spread and contaminate the lab. Consult your instructor for inspection and proper disposal.
- Do not use agar plates (solid media) or liquid media that appear to have bacterial growth / contamination. Consult your instructor for inspection and proper disposal.
- Immediately clean spills by flooding with disinfectant for a few minutes and wiping with appropriate absorbent material. For large spills, immediate inform your instructor.
- Always be respectful of your lab mates and their workspace.
- Avoid distracting and obstructing lab mates when carrying out an experiment for this can compromise their safety and the integrity of their experiments.
- Communicate questions or doubts about lab safety and proper technique at all times. Also, communicate when others are not adhering to lab safety rules and nonhazardous techniques.
- Always keep a mental note and written-record of your experiments. If you are lost or confused, do not hesitate to ask.
- Be aware of yourself and your surroundings at all time.
Aseptic or sterile technique is a central concept in microbiology and may be the most important part of working in a microbiology lab.
The goal of aseptic technique is to promote practices that 1) prevent the contamination of cultures, lab supplies and equipment with bacteria or fungi from the environment, and 2) prevent the contamination of individual working in the lab with potentially pathogenic bacteria.
Aseptic technique requires constant attention until it becomes second nature. Practice and strict adherence to these practices is key to making proper aseptic technique a habit; the health of individuals in the lab and the integrity of experiments depend on these principles. We will be learning aseptic technique in the lab though here are some general guidelines for practicing aseptic technique:
- Remember that bacteria are ubiquitous in the environment and our bodies.
- When possible, light a Bunsen burner or flame when working with bacteria – either as solid or liquid cultures. The Bunsen burner will create an umbrella of sterility by creating an upward airflow in your workspace and preventing debris from falling on your sterile materials. Work at a safe distance from your flame without risking burning yourself or anything in your workspace.
- Handle lab materials sensibly, avoiding unnecessary motions or contact with contaminated objects/surfaces or things that are not pertinent to the protocol at hand. Inoculating loops, sticks, toothpicks, pipette tips, serological pipettes, agar plates, media, and other previously-sterilized materials are packaged and stored under aseptic conditions and should remain as such for use. Should you inadvertently touch any items directly engaged in the protocol, the item is no longer sterile and must be re-sterilized or discarded (if the item is disposable).
- Keep in mind that Petri dish lids prevent particulate matter and airborne bacteria from contacting the plate surface, while allowing the diffusion of air around the edges. Whenever a plate lid is removed, it should be held over the plate as a shield. Do not place the lid face down on the bench top; avoid placing it on the bench all together. Do not leave plates uncovered. Do not walk around the room with an open plate.
- When working with cultures in test tubes, work diligently and as fast as possible. If Bunsen burners are available, pass the uncapped mouth of test tubes through the flame to prevent air from coming into the test tube. Keep tubes open for a minimum amount of time. While lids are removed, hold the tube at a slight angle so that airborne contaminants cannot fall into the open tube. Point tubes away from your face and body while working with them.
Working with Unknown Microbes
Based on ASM guidelines: Emmert et al. Biosafety guidelines for teaching laboratories. 2013. JMBE. 82, 14(1)
When working with environmental samples, we are always faced with unknown microorganisms. Based on what we know of soil bacteria, most will probably be harmless or nonpathogenic. These bacteria are treated as low-risk or biosafety-level 1 (BSL1) microorganisms. However, bacteria that are normally nonpathogenic may become pathogenic under unusual conditions or environments. Furthermore, we can never disregard the possibility of isolating a pathogen. These bacteria are treated as moderate- to high-risk or biosafety-levels 2 or 3 (BSL2/3), respectively. For these reasons, we should always treat bacterial isolates in the lab as potential pathogens.
Another risk of isolating unknowns is bacterial load. Every time we plate a soil sample, we are allowing individual bacterial cells to reproduce exponentially under optimal conditions. Colonies contain millions of cells, where only a small number may be necessary for infection. Students with a greater risk of infections – due to compromised immune systems or prolonged sickness – should inform their instructor.
The following guidelines, based on ASM “biosafety guidelines for teaching laboratories,” contain precautionary measures that we must take when working with unknown isolates:
- Treat all unknown bacterial isolates regardless of sample source (nature, household items, human skin) as potential pathogens or microorganisms that may need BSL2 containment, and in rare cases BSL3 containment.
- BSL1 labs may plate bacteria from environmental samples; however, these plates must remain covered and sealed or stored in secure location. Isolates must remain enclosed at all times, and only observed through the lid. After observation, plates must be decontaminated by autoclaving and properly discarded.
- BSL2 labs may subculture, or transfer, unknown bacterial isolates from the original medium (described above) to a fresh medium for further testing.
Labeling materials and supplies is a critically important aspect of science. With multiple plates, tubes, and samples, it is easy to become confused and lose track of things between and within experiments without the proper labeling practices. Here are a few guidelines with labeling:
- Always label the back of the petri dish – never the lid as the lid may become separated from the actual dish containing the growth medium (for example, if plates are dropped).
- Always add a date. Dates help orient the scientist as to when the experiment was performed and provide information about the quality of materials.
- Write as legibly as possible – others may need to find your materials or experiments.
- Label experiments with your lab table letter, your table number, your initials, and the bacteria and/or experiment and/or number of trials.