Collins Lab Blog

Reagent preparation

One important thing I learned in my graduate student days was the importance of tracking each reagent used in an assay. I was taught to keep a meticulous log recording the manufacturer, reagent grade quality, lot number, storage and preparation conditions for all reagents.

One episode in particular made a big impression on me in this regard. At the time, our lab was in the process of setting up an assay, the famous intra-Golgi transport assay created in the laboratory of Jim Rothman (Balch et al. (1984) Cell 39:405–416). Our lab had received a package containing all the reagents used for the assay. One of these reagents was non-fat dried milk. I was naively surprised that non-fat dried milk was included in the package, after all, one can visit any supermarket to get hold of this. But my advisor explained to me the importance of consistent sourcing and manufacturing for all components of a biological assay and impressed upon me that this meticulousness in attention to detail was a key underpinning of experimental success. Even in the case of humble reagents like non-fat dried milk.

Particularly important to know is how to handle each reagent or chemical after its arrival in the lab, where to store it, under what conditions and the length of the time period it could be used reliably before ordering a new batch. Although obviously critically important to the outcome of an experiment, this type of crucial information is mostly not recorded in the published literature and can make reproducing experiments between labs a challenge. One is mostly reliant on a combination protocols traditionally used by each particular lab (aka “lab lore”), together with any handling specifications provided by the supplier.

If you are fortunate, you may be able to accurately source a study in the literature that evaluates different protocols such as this,

which reports the stability of PMSF, a commonly used protease inhibitor, in aqueous buffered solutions. It is not uncommon for many graduate students to resort to internet searches to find reagent handling protocols which can be very problematic. With the advent of books like the Current Protocols series, the situation has improved somewhat, with standardized refereed information becoming more widely available to handle commonly used reagents.

So I was surprised the other day when I needed to make a solution of PMSF. In my experience, this chemical is always stored at RT. Searching the chemical shelves in my lab didn’t turn up any PMSF, however I was fortunate to locate some in another lab who generously shared some with me.

Only later did I realize why I couldn’t find this reagent in our lab. It was because our bottle was in the freezer!

I have no idea why this particular manufacturer, GoldBiotechnology, recommends storing PMSF at -20˚C.

No wonder it is a challenge for inexperienced experimentalists to negotiate their way through the vast and confusing array of information.

 

ETA

I emailed GoldBiotechnology to ask why they recommend storing PMSF at -20˚C and received this reply:

“When no data is available on the effect of long term storage at room temperature we default to storage desiccated at -20˚C.”

So here a chemical supplier is using a default recommendation. Because of the hygroscopic nature of PMSF and the fact that it is destroyed rapidly upon contact with water, it would probably be more practical to store the solid chemical at RT.

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