Tris Buffered Saline (TBS) - Histology

What is Tris Buffered Saline (TBS)?

Tris Buffered Saline (TBS) is a buffer solution commonly used in histology and molecular biology. It consists of Tris (tris(hydroxymethyl)aminomethane) and sodium chloride (NaCl) dissolved in water. TBS is used as a wash buffer in various experimental protocols, particularly in immunohistochemistry and western blotting.

Why is TBS used in Histology?

TBS is preferred in histology because it maintains a stable pH, typically around 7.4 to 8.0, which is crucial for preserving the antigenicity of proteins and the structural integrity of tissues. Unlike other buffers, TBS does not interfere with the binding of antibodies to their targets, making it ideal for immunostaining procedures.

How is TBS Prepared?

The preparation of TBS typically involves dissolving Tris and NaCl in distilled water. The desired pH is then adjusted using hydrochloric acid (HCl). A standard TBS recipe might include:

20 mM Tris
150 mM NaCl
pH adjusted to 7.4 using HCl

What are the Variants of TBS?

Various formulations of TBS exist to meet specific experimental needs. Common variants include:

Tris Buffered Saline with Tween 20 (TBST): Includes the detergent Tween 20 to reduce non-specific binding.
Tris Buffered Saline with EDTA (TBS-EDTA): Contains EDTA to chelate divalent cations, often used in antigen retrieval.
10x TBS: A concentrated stock solution that is diluted before use.

Applications of TBS in Histology

TBS is widely used in various histological techniques, including:

Immunohistochemistry (IHC): TBS is used as a wash buffer to remove excess antibodies and reduce background staining.
Western Blotting: TBS serves as a wash buffer to rinse membranes after antibody incubation.
In Situ Hybridization (ISH): TBS can be used to wash tissue sections to maintain isotonic conditions.

Advantages of Using TBS

Some of the key advantages of using TBS include:

Maintains stable pH, preserving protein structure and function.
Does not interfere with antibody-antigen interactions.
Compatible with a wide range of histological techniques.

Limitations and Considerations

Despite its advantages, there are some limitations and considerations when using TBS:

TBS without detergents may not be sufficient to reduce background staining in some protocols.
The presence of divalent cations in TBS can sometimes interfere with certain experiments; hence, TBS-EDTA might be preferred.
Proper pH adjustment is crucial; any deviation can affect experimental outcomes.

Conclusion

Tris Buffered Saline (TBS) is a versatile and essential buffer in histology, providing a stable environment for various protein-based assays. Its ability to maintain pH and compatibility with antibodies makes it indispensable for techniques such as immunohistochemistry and western blotting. Understanding its preparation, variants, and applications can significantly enhance the accuracy and reliability of histological experiments.