Small Particles Task Force: Minimization of Contamination – Best Practices (MMC)

This post is about how I prevent contamination in the lab during experiments and material preparation.

Contamination has been an issue in sample preparation despite efforts being made to minimize contamination. Current efforts include wearing cotton lab coats across all projects and the MMC, triple rinses with tap water followed by triple MilliQ water, working in chemical fume hoods or biosafety cabinets, keeping samples contained in containers while not in use, cleaning working spaces before use and before a new sample type is introduced, and washing gloves off before use.

Glassware Cleaning: Until recently, glassware was cleaned with filtered water and Isopropyl alcohol in a cleanroom. However, contamination was still an issue, despite improvements compared to non-cleanroom prepared glassware. Cleanroom preparation takes about half a day of work to prepare glassware that would typically be used in experiments, thus being a slow way of carrying out experiments. While no longer the go-to, protocols for filtering IPA for glassware cleaning can be found here. Additionally, filtering water for glassware cleaning or to be used elsewhere can be found here.

An easier way of cleaning glassware is to use 1% Alojet solution with tap water and rigorously shake and scrub the container, followed by 3x rinsing (or until detergent is visibly gone) under running tap water. This is then followed by rinsing 3x with MilliQ water (20% volume of the container for each rinse) followed by immediately capping or placing aluminium foil over your container.

Nalgene Filtering of Liquids:  A way of getting clean solutions while not going through cleanroom-level cleanliness rigor is to use cell culture grade consumables (tubes, dishes, bottles, etc.) that come pre-sterile and are virgin plastics. Through repeated use, shedding from these items was not seen. For this reason, many of the liquids prepared by the MMC are filtered using Nalgene high-volume filters (250 – 500 mL). PES Nalgene filters are used to filter MilliQ water and 10% KOH solution. Cellulose Nitride Nalgene filters can also be used for MilliQ water only. A protocol on how to use a Nalgene filter to filter your liquids can be found here.

These Nalgene Filters can be found by following this link. You may select from the variety of options that fit your needs.

Cleaning Work Area: Whenever I work on my microplastic research, I am always in a fume hood. Before putting in my supplies, I generously spray the entire fume hood bench space with 70% EtOH and wipe the bench top with a paper towel. I also do this after I am done working in the hood, too, ensuring that anything (like particles or solutions) is fully wiped clean before leaving. This helps prevent particles from sticking to the bench top, making it harder to dislodge later on.

Rinsing off gloves: Most laboratory gloves (including powder-free ones) have some release agent on them from the manufacturing process. This can get into samples if not removed. With every glove change, I spray my hands with 70% EtOH, rub my hands together like I am washing them, and then rinse off the EtOH under running water for 3-5 seconds. I then dry my gloves with a paper towel. If I have to handle multiple types of particles within the same experiment, I change gloves for each type.

Keeping Containers Closed: I often have jars, beakers, vials, and tubes in my fume hood while preparing samples and particles for the MMC. I keep everything closed with its lid/cap or with a sheet of aluminium foil until ready to be used, despite being in a fume hood. While airflow in the fume hood generally does a good job at preventing particles from settling onto things, the constant moving of arms and supplies can result in particles falling onto items (such as from clothing, skin, and other debris). This is especially important if there are dry powders being worked with, since even if you are highly thorough, things may still aerosolize and get into unwanted samples or containers.

Filtering Stains: 0.5 mL – 1 mL of the stock stains are filtered through 0.5 micron nanomembranes and stored as the working solution. I do not check every filtered stain individually, but do check the first sample I stain with a new batch of stain to ensure contaminates before not introduced to it based on the prestained condition of the sample.

More handling = more contamination: I avoid unnecessary handling (cleaning, baking, dipping, etc) whenever possible. For example, if I have to transfer liquid or powder from one container to another and the mass/volume does not matter, then I just pour from one to the other instead of using a pipettor or a scooper. I also avoid baking glassware and instead clean it with water before and after I use it and seal it. I find that more handling to make things cleaner, the worse the outcomes become due to increased exposure.