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pH Measurement Basics
TRIS buffers, which are often present in biological samples, can cause challenges in pH measurement. The interaction between classic silver-silver chloride electrolyte solutions and the key components of the TRIS base leads to junction blockage and shortened pH sensor lifespan. Selection of the appropriate reference system can allow improved pH sensor accuracy and extended lifetime.
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Any pH measurement requires two electrodes: a glass sensing electrode, and a reference electrode. Commonly, pH sensors in use today are so-called 2-in-1 sensors, meaning that both the sensing and reference electrodes are housed in the same sensor.
Figure 1: Reference and measuring sensor combined in a 2-in-1 sensor
The most commonly used reference system in pH measurement is silver/silver chloride. A conventional silver/silver chloride reference system is comprised of a silver wire coated with silver chloride and a reference electrolyte which is in contact with the solution of interest through a junction. For this construction, it is important that the reference electrolyte be saturated with silver chloride, so that the silver chloride is not depleted and stripped from the wire. In order to maintain a constant potential, the silver chloride must not be depleted, and the junction must allow the flow of ion-rich electrolyte into the sample.
Ordinary silverchloride sensors cannot be used for measuring TRIS samples. Many compounds are known to bind with silver ions to create large molecules or precipitates. In fact, silver chloride itself is one such precipitate, with limited solubility in aqueous solutions. Another such compound is formed between silver ions and TRIS molecules, commonly found in biological buffers.
Silver ions in a standard reference electrolyte solution flow into the sample of interest. If that sample contains a TRIS buffer, those two species will meet to form a stable, precipitating complex. Unfortunately, the point where this precipitate forms is typically the reference junction, which is a small ceramic frit with microscopic pores. This precipitation reaction clogs the pores in the junction, preventing electrolyte from flowing out. Once this process occurs, the interface between the sample solution and the reference system is eliminated, and the reference system can no longer generate a constant potential. This introduces unstable and erroneous pH readings.
Figure 2: Left to right: clean junction (white “dot”) in front of a black temperature probe and contaminated junction (black “dot”)