Tris buffer

Tris buffer (Tris(hydroxymethyl)aminomethane, also known as THAM or trometamol) is the most commonly used buffer in biological and biochemical research. Its popularity stems from its effective buffering range near physiological pH and its compatibility with many common laboratory applications.

🧪 Key Chemical Properties

• Chemical Formula and Weight: C₄H₁₁NO₃; molecular weight of 121.14 g/mol.

• Appearance: White, crystalline powder.

• Useful pH Range: 7.0–9.0.

• pKa: 8.1 at 25°C; 8.3 at 20°C; 7.82 at 37°C.

• Temperature Dependence: The pH of Tris buffer changes significantly with temperature. It increases by about 0.03 units for every 1°C decrease in temperature, and decreases similarly with a temperature increase. Important: Prepare your Tris buffer at the temperature at which it will be used.

• Concentration Dependence: The pH decreases by 0.03–0.05 units with each ten-fold dilution.

• Solubility: Highly soluble in water (saturation at 0°C is 2.40 M).

🔬 Common Applications

Tris buffer is a workhorse in many labs, with key uses including:

• Electrophoresis: A core component of TAE (Tris-acetate-EDTA) and TBE (Tris-borate-EDTA) buffers for nucleic acid agarose gel electrophoresis, as well as Laemmli buffer for SDS-PAGE protein separation.

• Nucleic Acid Storage: Used in TE buffer (10 mM Tris-HCl, 1 mM EDTA) to dissolve and protect DNA and RNA.

• Protein Analysis: Used in Western blotting (e.g., in TBST buffer) and as a component of RIPA lysis buffer for protein extraction.

• Other Uses: Found in cell culture media (at specific concentrations), diagnostic tests, and in the formulation of some pharmaceuticals like vaccines.

⚠️ Important Considerations & Limitations

Despite its widespread use, Tris has several limitations to be aware of:

• Enzyme Interference: The primary amine group can act as an inhibitor and has been reported to interfere with the activity of many enzymes. It is not suitable for use with the BCA or Bradford protein assays.

• Cell Toxicity: It can be toxic to some mammalian cells and can permeate cell membranes, making it unsuitable for many cell culture applications.

• Chemical Reactivity: It can inactivate DEPC (a common RNase inhibitor) and may form Schiff bases with aldehydes and ketones.

🧑‍🔬 Preparation and Forms

• Preparation: A common method to prepare a 1 M Tris solution is to dissolve 121.14 g of Tris base in ~800 mL of water, adjust to the desired pH with concentrated HCl, and then bring the volume to 1 liter.

• Available Forms: Tris is available as a free base (Tris base) and as its conjugate acid (Tris-HCl). For a buffer, these are often mixed, or a Tris-HCl solution is adjusted with a base. Pre-mixed, sterile solutions and "pHast Pack" powder blends are also commercially available for convenience