What Does "Fixed" Mean in Histology?
In the context of histology, the term "fixed" refers to the process of preserving tissue samples to maintain their structure and composition as close to the living state as possible. This is a crucial step in preparing tissue for subsequent microscopic examination and analysis.
It prevents autolysis, which is the self-digestion of cells by their own enzymes.
It halts bacterial decomposition.
It stabilizes the tissue's proteins and cellular structures.
It enhances the tissue's resistance to the subsequent processing steps, such as staining and sectioning.
Chemical Fixatives
Chemical fixatives work by cross-linking proteins and other biomolecules. Some common chemical fixatives include:
Formaldehyde: Often used in the form of formalin, it is one of the most widely used fixatives due to its ability to cross-link proteins effectively.
Glutaraldehyde: Another aldehyde fixative, it is commonly used for electron microscopy as it provides excellent preservation of ultrastructure.
Alcohols: Ethanol and methanol are used for rapid fixation but can cause tissue shrinkage.
Acetic Acid: Often combined with other fixatives to preserve nucleic acids.
Physical Fixatives
Physical methods, such as
cryopreservation, involve freezing the tissue rapidly to preserve its structure. This method is particularly useful for preserving enzyme activity and certain antigens.
How Is Fixation Performed?
The fixation process generally involves immersing the tissue sample in the fixative solution. The duration and temperature of fixation can vary depending on the tissue type and the fixative used. It is crucial to ensure that the tissue is adequately penetrated by the fixative to achieve uniform preservation.
Tissue Thickness: Thicker tissues require longer fixation times to allow the fixative to penetrate completely.
Fixative Concentration: Higher concentrations can achieve faster fixation but may cause excessive cross-linking, which can obscure cellular details.
Temperature: Increased temperature can accelerate fixation but may also increase the risk of tissue distortion.
pH: The pH of the fixative solution can affect the preservation of cellular components, particularly nucleic acids.
It can cause alterations in the tissue's morphology and biochemistry.
Certain antigens may be masked, making them difficult to detect in immunohistochemical studies.
Over-fixation can lead to excessive cross-linking, making the tissue brittle and difficult to section.
Conclusion
Fixation is a cornerstone in the field of histology, ensuring that tissue samples are preserved in a state that closely resembles their natural condition. While various fixatives and techniques are available, the choice depends on the specific requirements of the study and the type of tissue being examined. Understanding the principles and limitations of fixation can significantly enhance the quality of histological analyses.
