Choosing the right biological buffer for your experiment can be tricky. The buffer choosing process is like playing “Guess Who.” Like the game, you begin with a very broad list of buffers and slowly ask questions and research answers to narrow down your selection.
By now you know all about Good’s Buffers and are looking for more guidance to help with this little game. Well, in this article, I’ve cut to the chase – scrapping the talks about Good’s Buffers and getting right into the other things you should consider. For more details about choosing the right buffer, troubleshooting and other information about biological buffers, check out our Buffer User Guide. It doesn’t cost a thing and it is loaded with all the information you need.
Step 1: Determining pH for your experiment
In many experiments, you’ll need to change conditions in order to keep your proteins happy in solution. Let’s say, for example, you’re doing an experiment at pH 7.4, which mimics biological conditions, but your protein isn’t stable at this pH, you’ll need to optimize the situation. A good thing to keep in mind is that your target pH is usually going to be about one pH unit away (sometimes slightly farther) from the protein isoelectric point (pI). The pI is the pH where a given molecule or protein carries no net charge. When a protein is at its pI, it isn’t quite as soluble. Calculating pI on your own can be a bit of a challenge. But don’t worry; we live in the age of the Internet, and there are plenty of online calculators out there such as the one from ExPASy.
Step2: Considering Unwanted Buffer Interactions
Another important step in deciding which buffer to use is determining if there would be any negative buffer interactions. For example, if you’re working with an enzyme, your organic buffer might influence enzymatic function for better or worse.
Step 3: Asking the Right Questions
Just like the game, you have to ask the right questions to be certain you’re getting the right buffer. While we have several listed in the Buffer User Guide, I’ve highlighted a few great questions here:
1.Are there any metals in solution which a buffer might complex with?
2.Is your target protein an enzyme, and what class does it belong?
3.What are the ionic strength requirements for ligand binding, protein stability or electrophoresis?
4.What literature exists that tells me about interactions of my proposed buffer and the system I am using?
All of these questions, plus the others listed in our guide are going to further help you find the most appropriate buffer for your experiment.
Step 4: Considering Downstream Experiments
One stumbling block that often happens for researchers is that they’ll start with a buffer and find out that it won’t work for downstream applications. For instance, Tris has been known to negatively interfere with the Bradford assay. Other biological buffers can negatively impact mass spec. Unfortunately, after steps 1 and 2, you have probably narrowed down your buffer only to find out it might not work so well down the road. It’s like a nightmarish version of “Guess Who.” In this case, you’ll have to do some research in finding appropriate alternatives; however, now that you’ve narrowed down the characteristics you need, doing that research will be a little easier.
Step 5: Sourcing the Right Buffers
Great, now you know exactly what buffer you’re going to use. It’s time to source your buffers. For most researchers, bottom line and purity are basically the most important criteria when it comes to where to buy buffers. Sure you can go to the top name guys and know you’re going to get the right product, but you also know it won’t be for the right price, even if you’re making your own buffers. Instead, GoldBio has a growing catalog of high-quality powder buffers and the price is exceptional.
Not only does GoldBio offer great products, but the products are backed up with excellent support and content to help you with your experiment. It’s further evidence of a company you can trust.
Be sure to check out our Printable Buffers Selection Guides – Arranged by pH or by Family!
Buffer Selection Guide Arranged by pH. Print, and
hang at your bench today!
Buffer Selection Guide Arranged by Family. Print and
hang at your bench today!
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"To understand the universe is to understand math." My 8th grade
math teacher's quote meant nothing to me at the time. Then came
college, and the revelation that the adults in my past were right all
along. But since math feels less tangible, I fell for biology and have
found pure happiness behind my desk at GoldBio, learning, writing
and loving everything science.
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