MES is a biological buffer used to maintain stable pH in the mildly acidic range, roughly pH 5.5 to 6.7. Its full chemical name is 2-(N-morpholino)ethanesulfonic acid, and it has the molecular formula C₆H₁₃NO₄S. MES belongs to a family called Good’s buffers, a set of compounds specifically designed in the 1960s for use in biological and biochemical experiments where traditional buffers caused too many problems.
Why MES Exists: Good’s Buffers
Before Good’s buffers were developed, researchers relied on phosphate, citrate, and other inorganic buffers that often interfered with the very reactions they were trying to study. Norman Good and his team set out to create buffers that met a strict list of criteria: they had to be chemically inert, non-toxic, highly soluble in water, poorly soluble in organic solvents, and unable to absorb UV or visible light. Critically, they also needed pKa values between 6.0 and 8.0, covering the pH range where most biological processes happen.
MES fits all of these criteria. It is one of several Good’s buffers commonly used in labs today, alongside HEPES, TES, CHES, and Tricine. Each covers a slightly different pH window, so researchers pick the one that matches their experimental conditions.
Key Chemical Properties
The pKa of MES is 6.10 at 25°C, meaning it buffers most effectively right around that pH. A buffer works best within about one pH unit above and below its pKa, which is why MES has a useful range of 5.5 to 6.7. Outside that window, its ability to resist pH changes drops off quickly.
Temperature shifts the pKa slightly. According to NIST reference data, MES has a pKa of about 6.27 at 25°C at zero ionic strength, dropping to roughly 5.97 at body temperature (37°C). The Promega temperature coefficient for MES is approximately −0.11 pKa units per 10°C rise. In practical terms, if you prepare an MES buffer at room temperature and then use it in a warm incubator, the pH will drift slightly downward. For most applications this shift is small, but it’s worth checking pH at the temperature you’ll actually be working at.
Why Researchers Choose MES
One of the biggest advantages of MES is that it has negligible metal-binding affinity. Many biological experiments involve metal ions, whether intentionally (studying metalloenzymes, for instance) or as trace components in a growth medium. Buffers that grab onto metal ions can skew results by pulling metals away from the proteins or reactions being studied. MES, along with HEPES and CHES, avoids this problem, making it a reliable choice when metals are part of the picture.
MES also doesn’t absorb UV or visible light, so it won’t interfere with spectrophotometric measurements. And because it dissolves readily in water but poorly in organic solvents, it stays in the water phase of experiments and doesn’t cross cell membranes easily, keeping it out of the cell interior where it could cause unwanted effects.
Common Applications
MES shows up frequently in protein electrophoresis. In SDS-PAGE, MES and MOPS are the two standard running buffer options for use with certain gel systems. Because MES has a lower pKa than MOPS, it produces faster run times and gives better separation of smaller proteins (low molecular weight range). MOPS, by contrast, is better for resolving larger proteins. Choosing between them comes down to the size of the proteins you’re trying to separate.
Beyond electrophoresis, MES is widely used in plant biology and microbiology experiments that require a mildly acidic pH. Plant cell culture media, enzyme assays at sub-neutral pH, and crystallography setups all commonly call for MES. It also appears in some histological staining protocols and in buffers for column chromatography.
Limitations to Keep in Mind
MES is not completely inert in every situation. Research on the calcium-sensing protein synaptotagmin found that MES can weakly interact with highly basic protein domains, likely because the sulfonate group on MES carries a negative charge that attracts positively charged protein surfaces. For most experiments this interaction is too weak to matter, but in sensitive binding studies involving basic proteins, it’s worth considering.
Toxicity is generally low, but not zero. Studies on plant tissue culture found that MES becomes toxic at high concentrations. At standard working concentrations (typically 10 to 50 millimolar), it’s well tolerated by most cell types. Even so, some researchers have observed that organic buffers including MES can subtly alter the physiological behavior of cells, so appropriate controls are always important.
Finally, MES only covers the acidic side of the biological pH spectrum. If your experiment needs a pH near 7.0 or above, you’ll want HEPES (useful range 6.8 to 8.2), PIPES, or another buffer with a higher pKa.
Preparing an MES Buffer
Preparation is straightforward. You dissolve the desired amount of MES powder (or its sodium salt, or monohydrate form) in water to your target molarity. Common working concentrations range from 10 mM to 100 mM. MES dissolves as a free acid, so the starting solution will be acidic. You then adjust the pH upward to your target using sodium hydroxide (NaOH), adding it gradually while monitoring with a pH meter.
A few practical tips: always adjust pH at the temperature you plan to use the buffer, since MES’s pKa shifts with temperature. If you’re making a stock solution at a higher concentration (say, 0.5 M or 1 M), dilution will also change the pH slightly, so a final check after dilution is a good habit. Store MES solutions at room temperature or refrigerated. The powder itself is stable and inexpensive, which was one of the original design goals for Good’s buffers.