Staining Techniques, In this process, we stain very thinly sliced sections of prepared tissue so that different muscle types can be identified separately. Stein’s method is as follows:
Processing of Staining:
There are two ways to stain:
- Manual method
- Mechanical method
Manual Staining Procedure:
- Place the section fixed on the slide in Xylol for 30 minutes. Alternately in two separate beakers.
- Then keep it in 100% alcohol for 3 minutes.
- Then place in 85% alcohol for 2 minutes.
- Now keep in Methylated Spirit for 2 minutes.
- Now wash the slide in running water for about a minute.
- And then place in Hematoxylin for five to seven minutes.
- Now wash the slide for half a minute and place it in one percent acid alcohol for fifteen seconds.
- Wash in running water for half a minute.
- Dip in ammonia water two to three times until the tissue turns pale blue.
- Rinse with water.
- Immerse the slide in Eosin for three to five minutes.
- Wash with water for half a minute.
- Now dip two to three times first in 50% alcohol, then in 70% alcohol and then in 95% and finally in 100% alcohol.
- Finally clear it with Xylol and mount with Canada Balsam.
Tissue that will be visible under the microscope.
|Nucleus||Bright blue color|
|Muscles||Bright pink color|
|The inner part of the cell||Light pink color|
|Red blood cells||Orange-red color|
This method requires a machine (Automated Tissue processing machine). In which there are many glass beakers and put different chemicals in them and set the time for each step. Under this program, the machine rotates and eventually stains the tissue. It is a useful machine. It must be used where there is a rush of work.
Some important information:
- Dye is a compound that can color various fibers and muscles.
- Chromophores = They do not have the ability to give color to anyone while it is itself a colored compound.
- Autochromes = (they are acid or alkali) but when they are added with a chromophore, the chromophore has the ability to give color and is converted into a dye.
- A mordant is a compound that reduces the contact between the dye and the tissue and increases or facilitates the staining ability of the dye. Sometimes it is added to the dye and does not have to be added separately.
According to classification, they are divided into four categories.
Acidin stains only basic compounds such as Eosin
And it gives a red shade.
It colors only acid compounds eg fuchsine basic and gives a blue shade.
When these acids and bases are mixed, a tan is produced because both shades are mixed, so they show different colors, eg Leishman’s stain.
When specific types of muscle fibers or cell interiors are to be stained, a special type of machine is made, eg PAS stain, etc.