Under the microscope: Buffered Formalin (NBF)

Under the microscope: Buffered Formalin (NBF)

Buffered formalin, also known as neutral buffered formalin (NBF), is a specific formulation of formalin that includes buffering agents. In histology, NBF is commonly used as a fixative for tissue samples. The buffering agents added to formalin helps to maintain a stable pH level, which is important for preserving the tissue’s cellular structure.

What Is Buffered Formalin (NBF) Used For?

Here are some key uses and benefits of buffered formalin in histology:


  1. pH stabilization: The buffering agents in NBF help maintain a constant and optimal pH level, usually around 7.0. This pH range is ideal for preserving tissue structures and minimizing the formation of formalin-induced artefacts.
  2. Enhanced tissue preservation: The stable pH provided by buffered formalin prevents excessive acidification or alkalization of tissue samples during the fixation process. This ensures better preservation of cellular components and minimizes degradation.
  3. Consistent and reliable results: Using buffered formalin helps standardize the fixation process, leading to more consistent and reproducible results in histological staining and analysis.
  4. Compatibility with downstream processing: Buffered formalin-fixed tissues are compatible with a wide range of tissue processing techniques, such as dehydration, clearing, embedding in paraffin, and sectioning for microscopic examination.
  5. Long-term storage: Like regular formalin, buffered formalin allows for long-term storage of fixed tissues without significant degradation, facilitating archival and future reference.


What Makes A Good Buffered Fomalin (NBF)?

A good buffered formalin in histology should possess several key characteristics to ensure effective tissue fixation and preservation. Here are some important factors that contribute to making a good buffered formalin:


01. Correct formalin concentration

The ideal concentration for buffered formalin in histology is typically 10% formalin. This means that the solution contains 10% formaldehyde gas dissolved in water, with the remaining 90% being water and buffering agents. This formulation helps to achieve optimal tissue fixation and preservation while minimizing the risk of formalin-induced artifacts.

Using a 10% formalin solution provides a good balance between effective fixation and tissue penetration. Higher concentrations of formalin can result in tissue surface fixation but may cause inadequate penetration into deeper tissues, leading to incomplete preservation. On the other hand, lower concentrations might not provide sufficient cross-linking of proteins and may lead to poor tissue preservation.


02. Buffering agents

The buffered formalin should contain buffering agents that help maintain a stable pH, typically around 7.0. This pH level ensures that the formalin fixation process is consistent and minimizes tissue damage or artifacts caused by pH fluctuations.


03. Rapid tissue penetration

A good buffered formalin should have the ability to rapidly penetrate tissues, ensuring even and efficient fixation throughout the sample. Rapid penetration helps prevent autolysis and decomposition of tissues and leads to better preservation.


04. Minimal tissue shrinkage

The ideal buffered formalin should result in minimal tissue shrinkage during the fixation process. Excessive shrinkage can distort tissue structures and lead to inaccurate microscopic interpretation.


05. Preservation of cellular morphology

The primary goal of tissue fixation is to preserve the cellular morphology and architecture as close to the living state as possible. A good buffered formalin should maintain cellular structures and prevent cellular distortion.


06. Compatibility with downstream processing

The fixed tissues should be compatible with subsequent histological processing steps, such as dehydration, clearing, embedding, sectioning, and staining. Proper fixation ensures that tissues maintain their integrity throughout these steps.


07. Long-term stability

The fixed tissues should be stable for long-term storage without significant degradation or loss of cellular details. This is crucial for archiving samples and future reference.


08. Safety

Buffered formalin is a hazardous chemical and should be handled with appropriate safety precautions. A good formulation should be labeled properly with clear safety instructions and handling guidelines. Some companies offer a prefilled buffered formalin, reducing the risks associated with using formalin.

responsible suppliers of buffered formalin should also be able to provide formalin control products or spill kits that contain items such as absorption pads, formalin absorption granules and formalin neutralizer to ensure the safe clean up and disposal of spilled formalin.


09. Quality control

Manufacturers should ensure strict quality control measures during the production of buffered formalin to maintain its efficacy and consistency.

It’s important to note that while buffered formalin is commonly used in histology, variations in protocols, tissue types, and research objectives may require adjustments in the formalin formulation or fixation techniques to achieve the best possible results. Researchers and pathologists should always optimize their protocols based on specific experimental needs and adhere to proper safety guidelines when working with formalin.