Tissue Preparation for Clinical Analysis

Tissue preparation is critical for the success of clinical analysis. If any step is missing or has been performed erroneously in the process of tissue preparation, the validity and reliability of the results from the clinical analysis done on the histological sample are immediately compromised.

Steps in Tissue Preparation for Clinical Analysis
Tissue specimens that have only been recently retrieved often appear spongy and transparent. They are vital sources of information in numerous clinical applications. Due to their essential makeup, however, tissue specimens must first undergo a careful process of preparation before they can be placed under a microscope for viewing.

There are four primary steps in tissue preparation and they are fixation, embedding, sectioning, and staining. Each step may produce artifacts. An artifact is anything that causes distortion in how a sample is seen. Artifacts may occur accidentally, inevitably, or by design. People can reduce chances of having artifacts present by using various techniques, but none of them are perfect. And as sometimes the occurrence of artifacts cannot be prevented, researchers have just learned to accept their presence as a normal part of certain tissue specimens.

The most typical process of tissue preparation would start with fixating the specimen using buffered formaldehyde, paraffin for embedding, slicing the sample with approximately 5 micrometers in thickness, and utilizing hematoxylin and eosin for staining.

Step 1 Fixation is the process of preserving and hardening tissue samples to make them retain their original condition as closely as possible. Various chemicals may be used for fixing tissue specimens. These are called fixing agents. They may be additive or non-additive, depending if they can combine with the tissue specimen’s proteins or not. They may also be coagulant or non-coagulant, depending on their ability to precipitate the tissue’s proteins.

Fixation must be performed quickly and this is made possible either by immersion or perfusion. In immersion, small samples are required but obtaining them through mechanical means could cause damage to the specimen. In perfusion, the fixative is injected directly into a live tissue specimen’s circulatory system but this is, of course, unfeasible for human patients.

There is another option besides fixation for retaining the original condition of a tissue specimen and that’s freezing. Frozen tissue specimens are immediately sectioned. Freezing may not be able to retain as much of a specimen’s original condition as fixation could but it is definitely a quicker process and can be immediately observed and yield preliminary results.

Step 2 After fixation, the tissue specimen must be embedded in a solid object. In the embedding process, solvents like xylene and alcohol replace tissue water. Epoxy solution or paraffin is used next and allowed to harden.

Step 3 A microtome is used to produce thin sections of tissue samples. The thickness required from a thin section depends largely on the clinical application it will be used for and the type of microscope utilized for viewing it. Several artifacts commonly occur in this stage of the process. Wrinkles are perhaps the most common and difficult to avoid. Tiny structures in tissue specimens may also disappear or appear in the wrong places when thin sections haven’t been properly sliced off. Chatter and scratches are also other typical artifacts. Chatter refers to visible records of knife vibration while scratches are caused by dirt or any other flaw found on the cutting edge.

Step 4 Since tissue specimens are generally transparent, they need to be stained properly to make them visible even under bright field illumination. Staining is also occasionally used to make certain parts of the sample distinguishable. As staining causes the color of the sample to change, color therefore must not be used as a tool for recognizing parts and making observations.

Using a Clinical Microscope
Proper use of a clinical microscope is the next important thing to focus on once you’ve correctly prepared your histological sample. While the basic functions of clinical microscopes remain the same among different models, higher end and custom-designed units may possess special and unique features that make them more suitable to a particular clinical application.

Make sure that your clinical microscope is placed on an even and firm surface before plugging it in. After switching it on and activating the light source, start adjusting from low to high power and look for the right eyepiece and objective to use for your sample. If you wish to save the images or even obtain video footage for live cells, connect the clinical microscope to a digital camera or video-recording device.

Lastly, remember to take down your observations immediately to prevent any loss of data.

Get to know more about clinical microscopy through http://www.clinical-microscope.com