Have you ever needed an enzyme but not had an ice bucket? Have you been tempted to just grab the enzyme out of the freezer, take what you need and quickly return the tube to the freezer? Do enzymes really need to be kept on ice all of the time? I would say, Yes!
Enzymes used in a research laboratory range from restriction enzymes used for plasmid digests, reverse transcriptases for cDNA synthesis reactions to polymerases used for PCR. Like many laboratory reagents enzymes are expensive! It is good laboratory housekeeping to maintain their longevity through proper storage and use. This doesn’t just make good economics, it also ensures the quality of your experiments. And it will keep you in the good graces of your labmates and PI!
Enzymes need to be stored and used at their optimum temperatures. Enzymes are generally stored in glycerol at -20C. This prevents them from freezing completely, which causes protein denaturation and results in a loss of activity. Enzymes will also denature if they are warmed up, repeatedly exposed to temperature fluctuations, or stored at room temperature for extended periods of time. Therefore, enzymes need to be kept cold at all times, even if you are just pipetting out of them quickly.
A little organization will help you set up your reactions without exposing enzymes to warmer temperatures. Make sure you have everything you need on hand prior to starting. If you are running an experiment that requires a programmed machine, make sure you book the machine and give yourself plenty of time to get everything set up. Program the machine prior to setting up your reactions. It is good practice to set up your reaction(s) by adding all other components first then adding your enzyme last. This is particularly important when working with Taq for PCR reactions; the polymerase can start working immediately so it is important to add the enzyme and get your samples into the machine quickly. Keep your enzymes in the freezer until just before addition. Bring them over to your bench in an ice bucket or enzyme cooler and return them to the freezer immediately after using.
So, to keep your enzymes happy, store them in the freezer and keep them on ice when working on the bench.
Human bronchial epithelial cells (HBECs) are a challenge to culture. As highly specialised cells that exist in carefully ordered multi-layered structures, they are especially fickle and finding optimum conditions to keep them happy is tricky. The cultures are also extremely sensitive to tiny changes in routine or environment. However, there are certain basic principles that […]
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