Question

Describe the most commonly used histological processes from specimen collection to H & E staining.

About 1000 words

Answer 1

Tissues from the body taken for diagnosis of disease must be processed in the histology labs to produce microscopic slides that are viewed under the microscope by pathologist.This is the histological process where tissue or specimens removed by surgey and proceed for making the microscopic slide for diagnosis of disease and histological study.So various steps are involved in this process. These are below explained.

1.Specimen Accessioning - For identification purpose specimens are accessioned by giving them number and keep lists of patient information and history alongwith description of the site of origin of sample.

2.Gross examination - Describing the specimen and placing all parts into a small plastic cassette .and it is being processed to a paraffin block.The cassettes are placed into a fixative.When malignancy is suspected the specimen is covered with ink in order to mark the margins.

3.Fixation - types of fixatives

The purpose of fixation is to preserve the tissues permanently.There are five major group of fixatives-

Aldehydes

Mercurials

Alcohols

Oxidizing agents-Potassium permanganate,dichromate and osmium tetroxide

Picrates

Many factors like Buffering,Penetration,Volume,Temperature,Concentration,Time interval and pH affects the Fixation process.Fixation is best carried out at neutral pH in the range of 6-8.

Formalin is used for all routine surgical pathology and autopsy(after death) tissues during H & E staining.

4.Tissue processing - The technique of getting fixed tissue into paraffin is called tissue processing.The main steps in processing are dehydration and clearing.Dehydration usually done by series of alcohols.For the removal of dehydrant commonest clearing agent is xylene.The tissue is infiltrated with the embedding agent always paraffin.A product called paraplast contains added plasticizer that make the paraffin block easier for some technicians to cut.A vaccum can be applied in the tissue processor to assist penetration of the embedding agent

5.Sectioning -Once the tissue has been fixed,it must be processed into a thin microscopic sections.Sections are 3 to 10 micons in size. by use of microtome

6.Frozen sections - Frozen sections are performed with an instrument called a cryostat.It is a refrigerated box temperature about -20 to -30 Celsius.

7.Staining - H & E(Hematoxylin and Eosin).it will not directly stain the tissue.It needs a mordant.

8.Coverslipping - Stained section must be covered by thin piece of plastic or glass to protect the tissue from being scratched.

We can understand these steps by below picture-