Understanding The Science Behind Tissue And Dealcoholization
While seemingly unrelated, tissue and dealcoholization, are two fascinating fields of study that delve into the intricate world of biology and chemistry.
Dealcoholization doesn’t only concern the production of low-alcoholic drinks and non-alcoholic beverages. It can also touch on medicine when we talk about dealcoholization of tissue, which is a part of tissue processing.
The science behind tissue and dealcoholization sheds light on the innovative techniques and principles that underpin these areas of research.
In the context of beverage production, dealcoholizationrefers to the process of removing or reducing the alcohol content in alcoholic beverages to create non-alcoholic or low-alcohol versions of these drinks.
This process is primarily done to meet the needs of consumers who either want to abstain from alcohol or reduce their alcohol intake for various reasons, such as:
- personal preferences
The goal is to create a non-alcoholic or low-alcohol beverage that closely mimics the sensory characteristics of the original alcoholic version while complying with legal regulations regarding alcohol content in beverages.
Dealcoholization methodscan vary depending on the type of beverage and the desired alcohol content, but some common techniques include:
- heat evaporation
- reverse osmosis
- reverse osmosis
Dealcoholized beerand wine have become increasingly popular in recent years. Several breweries and wineries now offer a range of non-alcoholic or low-alcohol options to cater to a broader customer base.
As for tissue and dealcoholization, which is part of tissue processing, the focus is on pertinent medical use and on the role that they play in research.
Dealcoholization of tissue is an important step in tissue processing, particularly in the field of histology (study of tissues) and pathology (study and diagnosis of diseases).
Tissue processing is the series of steps that are carried out to prepare biological tissue samples for microscopic examination.
Tissue and dealcoholization are involved in transitioning tissue samples from fixative solutions (usually containing alcohol) to a form suitable for embedding in paraffin wax or other embedding media and subsequent sectioning for microscopic analysis.
Here’s how dealcoholization of tissue fits into the broader context of tissue processing:
a. Tissue Fixation
The first step in tissue processing is tissue fixation.
Tissue samples are preserved in fixatives, which typically contain:
- formalin (formaldehyde)
- other chemicals
Fixation helps to stabilize cellular structures and prevent tissue decomposition.
After fixation, tissues are dehydrated to remove water from the tissue.
This is done through a series of graded alcohol solutions, typically starting with a low concentration of alcohol and gradually increasing the concentration.
Dehydration is a critical step to ensure that the tissue is ready for embedding, as embedding media (e.g., paraffin wax and alcohol) don’t mix well. So, tissue and dealcoholization happen.
Following dehydration, the tissue needs to be “cleared.”
This involves the removal of the alcohol from the tissue, which is essential before embedding in paraffin wax.
The process of clearing essentially means replacing alcohol with a substance that is miscible - can be mixed - with paraffin wax (e.g., xylene or a xylene substitute).
This step is important to ensure that the tissue isn’t compromised when it comes into contact with the embedding medium.
d. Infiltration and Embedding
After clearing, the tissue is infiltrated with paraffin wax or another embedding medium, ensuring that it penetrates the tissue fully.
This step allows the tissue to be embedded in a solid block, making it easier to section.
Once the tissue is embedded, it is allowed to solidify, and thin sections can be cut for microscopic analysis.
This step is crucial for the successful transition of tissue from the dehydrated state to the embedded state.
Without proper dealcoholization, the tissue may not interact well with the embedding medium, leading to poor-quality tissue sections that are not suitable for microscopic examination.
Therefore, tissue and dealcoholization are vital in tissue processing, particularly during the clearing step.
The dealcoholization of tissue ensures that tissue samples are appropriately prepared for embedding and sectioning, ultimately enabling the analysis of cellular structures and pathological features under a microscope.
Perhaps the most popular clearing agent when we talk about tissue and dealcoholization is xylene, a colorless, sweet-smelling liquid that is highly flammable.
In addition to xylene and ethanol, other clearing agents that may be used in the dealcoholization of tissue include:
- cedarwood oil
- propylene oxide
According to the study Histology without Xylene, published by Annals of Diagnostic Pathology in 2009, the following clearing agents are good xylene substitutes:
- isopropanol, aka isopropyl alcohol
- blend of isopropyl alcohol and molten paraffin
- blend of isopropyl alcohol and mineral oil
In 2018, the Journal of Laboratory Physicians published Alternative to Xylene as a Clearing Agent in Histopathology. This study endorsed the use of UltraClear.
Compared to xylene, this clearing agent, as attested by the study, is:
- “less toxic”
- “less toxic”
- more environment-friendly
The only downside? UltraClear is more expensive than xylene.
The choice of dehydrating agent for tissue and dealcoholization may vary depending on the specific tissue type, the subsequent processing steps, and the goals of the tissue preparation process.
When discussing tissue and dealcoholization, alcohol will never be missed.
In the context of tissue processing, alcohol is used to remove water from biological tissues in a controlled manner.
The main role of alcohol is to replace the water in the tissue with a substance that can be easily infiltrated with paraffin or other embedding media, allowing for the creation of thin tissue sections for examination under a microscope.
Here are some key roles and functions of alcohol in tissue processing, which involves tissue and dealcoholization:
a. Water Removal
The primary function of alcohol is to remove water from the tissue sample.
Water can interfere with subsequent processing steps, and alcohol helps ensure that the tissue is properly dehydrated before further processing.
b. Preservation of Tissue Structure
Alcohol is used to preserve the tissue’s structural integrity and prevent shrinkage or distortion during the removal of water.
c. Facilitation of Infiltration
Dehydration makes the tissue more compatible with the embedding medium (e.g., paraffin wax) used in the final steps of tissue preparation.
Proper dehydration ensures that the tissue can be infiltrated with the embedding medium, allowing it to be sectioned into thin slices for microscopic examination.
Alcohol has a clearing function, helping to make the tissue more transparent and improving the visibility of cellular structures under the microscope.
Proper dehydration is a crucial step in histology and pathology to ensure that the tissue sections are of high quality and that cellular structures are preserved for accurate analysis and diagnosis.
To make it proper, it necessitates the use of alcohol.
Speaking of alcohol, according to a document titled Method of Processing Tissue Specimens and Dehydrant Solvent for Use Therein, the one usually used in tissue and dealcoholization is ethyl alcohol.
A recommended substitute will be a combination of isopropyl alcoholand methyl alcohol.
They should be mixed according to this amount (by volume):
- isopropyl alcohol - between 55 and 70 percent
- methyl alcohol - between 30 and 45 percent
An ideal blend (again, by volume) will be 60 percent isopropyl alcohol and 40 percent methyl alcohol.
In tissue processing for research or medical purposes, alcohol - particularly ethanol - is commonly used as a fixative and dehydrating agent.
Ethanol is a widely used dehydrating agent because it is:
- relatively safe
- readily available
- can effectively remove water from tissues without causing significant damage
However, there are alternative methods and reagents available for tissue and dealcoholization. The choice of alternative will depend on the specific requirements of the research or medical application.
Some alternatives to alcohol in tissue processing include:
Formalin is a commonly used fixative in histology and pathology.
It preserves tissues by cross-linking proteins and is an alternative to alcohol for fixing tissues.
b. Bouin’s Solution
French biologist (biological scientist) and endocrinologist Pol André Bouin, M.D. (1870-1962) invented this solution.
Bouin’s solution is a fixative that is a mixture of:
- picric acid
- acetic acid
It’s often used in histology and provides excellent tissue fixation.
c. Carnoy’s Solution
Belgian biologist Jean-Baptiste Carnoy (1836-1899), who also happened to be a Roman Catholic priest, created this solution.
Carnoy’s solution is a fixative made from a mixture of:
- glacial acetic acid
It’s particularly useful for preserving cell morphology and is commonly used for cytological preparations.
Glutaraldehyde is another fixative that can be used as an alternative to alcohol. It’s often used for electron microscopy to better preserve ultrastructural details.
e. Acetic Acid-Acetone Solution
A mixture of acetic acid and acetone can be used as a fixative and dehydrating agent, which is particularly useful for certain specialized staining techniques.
f. PFA (Paraformaldehyde)
Paraformaldehyde is a formaldehyde-based fixative often used in research applications, such as:
- fluorescence microscopy
g. Tissue Embedding Media
Instead of alcohol-based clearing agents, you can use various embedding media to infiltrate and embed tissues for sectioning and microscopy.
These embedding media include:
- paraffin wax
- glycol methacrylate
h. Ethylene Glycol
Ethylene glycol can be used as a dehydration agent in tissue processing, especially for frozen tissue sections.
It’s often used to replace ethanol or isopropanol.
i. Liquid Carbon Dioxide (CO2)
Liquid CO2 can be used for rapid freezing of tissues and is suitable for cryosectioning (frozen tissue sectioning), aka frozen section biopsy.
It’s an alternative to alcohol-based freezing methods.
j. Dry Ice (Solid CO2)
Dry ice can also be used for freezing tissues for cryosectioning, making it a suitable alternative to alcohol.
The choice of the alternative method or reagent will depend on the following:
- type of tissue being processed
- specific needs of the tissue processing protocol
- downstream applications or downstream processing (DSP)
DSP - when biological materials are taken from tissues (plant or animal tissues) - includes:
- molecular analysis
- electron microscopy
Consult with experienced histologists or researchers to determine the most appropriate alternative for alcohol in tissue and dealcoholization.
Yes, tissue absorbs alcohol.
In fact, according to an article by Bowling Green State University, when an individual consumes an alcoholic drink, the alcohol will enter “all tissues of the body.”
An estimated 68 percent of the tissues in the body of an adult male will absorb alcohol.
Alcohol won’t penetrate the fats and the bones, though.
Tissue and dealcoholization are primarily involved in the “clearing” step of tissue processing, where alcohol is removed from the tissue and replaced with a substance like xylene.
In tissue processing and in the dealcoholization of tissue, biology intersects with chemistry and vice versa.
By removing alcohol from tissues, researchers can unlock new insights into the structure, function, and interactions of biological molecules without the interference of alcohol’s solvent properties.
The science behind tissue and dealcoholization represents a promising avenue for advancing our understanding of various biological processes and improving medical research and clinical applications.