Tissue Processing Steps, Dehydration, clearing, impregnation, embedding histopathology lab

Tissue processing is an important procedure in the histopathology laboratory. It is used to preserve the anatomy of the tissue.

In order to cut the fine section of tissue, we need suitable hardness and consistency in the tissue.

Therefore these properties can be imparted by the infiltration, by the paraffin wax, celloidin, and by resins. Some tissue needs freezing.

What is the Principle of human tissue processing in histopathology?

It is a crucial procedure in histopathology. In order to cut the perfect section of different tissue-like, kidneys, lungs, tc. We must have hardness in tissue before presenting to the kife if microtome.

This hardness is achieved by the infiltrating agents. There are four stages of tissue processing.

What are the Tissue Processing Steps in histopathology laboratory?

It is a long procedure, it takes 24 hours. It can perform in two ways.

  1. Manually procedure. Humans are involved in this procedure.
  2. Automatically procedure. It is totally automated. The processor has all kinds of jars with their reagents.

Tissue Processing Steps, Dehydration, clearing, impregnation, embedding

What is Tissue Dehydration by Alcohol?

First of all, we need to remove water from the tissue. It  Dehydration is the first step. In this step, we use increasing strength of alcohol like 70%, 90%, and absolute alcohol for one hour.

Note: Dehydration time depends on three main factors.

  1. Size of the different specimens.
  2. Fixative
  3. Types of tissue

For the routine section (7 mm thick section), the following scheme is used.

  • First place in the 70% alcohol for one hour
  • Second, place in 90% alcohol (rectified spirit). Two changes for two hours.
  • Final, absolute alcohol. two changes for two hours.

Note:  Agitation is necessary for conventional tissue processing.  If there is no agitation, dehydration will take time than two hours. The dehydrating agent should be 50 to 100 times more as compared to tissue size.

Tissue Clearing by Xylene

It is an alcohol removal process. After dehydration steps in tissue processing, we need to clear alcohol by clearing agents. In this stage, alcohol is replaced with xylene this process is called clearing.

What are the Clearing agents in tissue processing in histo lab?

  • Chloroform
  • Xylene
  • Benzene
  • Carbone Tetrachloride
  • Toluene
  • Cedarwood oil

NOTE: Xylene is the most common cleaning agent.

A small piece of tissue is cleared in 1/2 or in 1 hour. But if the tissue is thick 5 mm or more than 5 mm, then it will take two to three hours.

Cedarwood oil is an excellent cleaning agent. However, it is slow in the clearing. It takes extra time.

Tissue Impregnation with Wax

When tissue is cleared by xylene or other clearing agents.  The next step is tissue processing embedding It occurs at the melting temperature of wax (55 to 56 centigrade).

Note: The volume of the wax should be 20 to 30 times more than tissue volume.

For better impregnation, 3 to 4 jars are necessary. Impregnation time depends on three factors.

  1. Tissue size
  2. Type of tissue
  3. Clearing agents

If xylene is used as a clearing agent, then it is easy to remove from the tissue. One to two changes of wax are sufficient.

How many types of wax are used for tissue impregnation?

There are the following types of wax used for impregnation.

  1. Paraffin Wax: It is used in the routine. It has hard consistency so 3 to 4 micro thickness sections can be cut.
  2. Water Soluble wax: It has a benefit, tissue directly placed in it without dehydration and clearing steps. BUT it has a disadvantage also, section fragmentation takes place in the floating bath.

What is the Importance of human tissue processing in histopathology laboratory?

It is an important procedure in the histopathology laboratory. In this laboratory, we deal with many organs like kidneys, lungs, and many others. Following are the key importance

  • Tissue Anatomy preservice
  • Can cut fine sections after processing

What is the purpose of tissue processing?

  1. It is used to preserve the anatomy of the sample like kidneys, lungs, pancreas, appendix same as they remove from the body.
  2. After tissue processing, we can cut fine sections from the organs for the histological study.

What are the 3 main steps of tissue processing in histopathology?

It has three basic steps as

  • Dehydration
  • Clearing
  • Embedding

What is the most important step in tissue processing?

Each stage of it is essential, but dehydration is the key step in it. If tissue is partially dehydrated, then other steps will be useless. We will be unable for a fine cut, and its anatomy will be destroyed during other stages.

Which kindes of tissues are used for tissue processing?

Following are mostly used in processing

  • Lungs
  • Kidney
  • Appendix
  • Skin
  • Uterus
  • Testes
  • Breast

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