I usually get to the lab quite early, in order to get a head start on my experiments before going to the usual BSURF meeting. Many of the experiments we run, such as Western Blots and RT-PCRs, require a lot of preparation and busy work (including a ridiculous amount of pipetting). A procedure can take up most of a day, if not multiple days, and you cannot simply stop and start at whatever point you like. There are set points in the procedure that you have to reach before calling it a day, and getting started early is therefore crucial, if you want to avoid having to stay late. For example, the membrane in a Western Blot has to be washed overnight in primary antibody, so that it can be developed the following day. Failing to set up a primary antibody wash before leaving would mean waiting a whole additional day for your results. For other procedures, the intermediates we work with are easily degraded, and must be converted to something else before being stored. RNA is particularly fragile, and after being isolated from cells must quickly be converted to cDNA. Therefore, you must perform the necessary reverse transcription reaction before finishing up for the day.
Another regular task is cell maintenance. After several days of growth, cells become overly “confluent” – crowded on their plate – and their growth media becomes depleted. In order to keep our cancer cell lines growing and healthy, we have to “passage” them every few days. This involves taking a fraction of the cells that were growing in culture and transferring them to a fresh plate with new growth media. A substantial amount of work goes into simply ensuring that you have cells to perform experiments on, let alone performing the experiments themselves!
This past week has been particularly busy, as I have been doing a time course of GNF5 (ABL kinase inhibitor) treatment – essentially, I treat cells with the drug at a number of different time points over a period of 72 hours. This has involved coming to lab several times a day, often at odd hours, ranging from 7AM to 11PM. The LSRC is a somewhat eerie building that late at night!
An entire experiment, from plating cells out for a drug treatment to producing presentable data by developing your Western Blot or analyzing your RT-PCR, takes at least several days. It is therefore important to enjoy the procedures themselves, and not only be in it for the results – otherwise you are likely to find yourself bored and frustrated most of the time.