Histology slide : Summary of tissue preparation with mcq

 

Histology slide 

Tissue preparation

 Under the light microscope, tissues are examined via a light beam that is transmitted through the tissue. But the tissue and organs are usually too thick for light to pass through them. For this region tissues have to sectioned to obtain thin, translucent sections. Before making the histological slide tissue has to pass though some preparation processes. Following are the steps of tissue preparations of histological slide.

Steps of tissue preparation for study  

1. Selecting parts of tissue: part of an organ selected and cut. The best thickness are 3-5mm
2. Fixation: The process of avoiding tissue damage from bacteria or autolysis(cell digestion by cells own enzymes) by the help of some chemicals (fixatives) is called fixative. Name of fixatives: Formaline: Formaldehyde  is a gaseous substance but 40% of it soluble in water. In tissue prepation we uses  :10% formaldehyde(CH20) solution in water.
• Uses of formalin: used as a disinfectant or to preserve biological specimens.
•  Health hazards: short time exposure:  irritation of eye,skin, mucosa of nose and throat, it is  carcinogenic substance  but not teratogenic.
• Precurtion during use of formalin :
•  Exposure to mild formalin for 8 hrs /per day or 40 hrs in a week is safe. Formalin is not absorbed through skin so mild formalin can be handling for short time. After handling wash hand with soap and plenty of water for few min. For long term handling use gloves. Contamination of eye with formalin: wash eye with water for 15 min and consult with ophthalmologist.

Dehydration : after fixation, tissue must be dehydrate by graded series of ethanol and water, usually from the 70% to 100% 

Clearing : after dehydration, tissue must be clean with xylene.

3. Infiltration: after clearing  tissue must be infiltrate with paraffin.

 What is paraffin: for infiltration, tissue must place into a container containing liquid paraffin and place it  in an incubator at the temperature of 58-60 C  

4. After that tissue must be place in a cash for embedding or block formation
5. then sectioning with microtome : now block of the tissue cut by microtome ( a cutting instrument ) 
6. Staining: unfortunately tissues are colourless can not be observe in microscope. So need to be stain. For staining rehydration is necessary. Rehydration is done by graded ethanol and water from 100% to 70%.
7. In routine staining hematoxilin and eosin

Name of the staining and its components:

Basophilic : Tissue components that stain more with basic dye is called basophilic

Acidophilia : tissue components that stain more with acidic dye is called acidophilic

Colour of different components of tissue:

• nucleus light blue to  dark blue
• Cytoplasm and extra-cellular matrix  light pink to red 
7. Dehydration: after staining tissue need to be dehydrate again for long preservation.
8. Clearing: after dehydration tissue need to be clean with xylene. 
9. Amounting: DPX. This chemical place above the tissue and covered by coverslip for long term use.
10. Now a histological slide is ready for study.
11. (Write T for True and F for False)

12. 1.        Name the fixative which is used in histology

    i)                     40% formaldehyde

    ii)                   xylene

    iii)                 paraffin

    iv)                  osmium tetroxide

    v)                   DPX

    2.        Following are the disadvantage of formaline ?

    i)                     preservation of tissue

    ii)                   irritant of skin, mucosa of nose and throat

    iii)                 teratogenic( harmful for fetus)

    iv)                  carcinogenic

    v)                   disinfected

    3.        Following is/are the clearing agent/agents

    i)                     paraffin

    ii)                   formaline

    iii)                 alcohol

    iv)                  xylene

    v)                   eosin

    4.        Following instrument is/are used for cutting paraffin block.

    i)                     slide

    ii)                   knife

    iii)                 coverslip

    iv)                  microtome

    v)                   oven

    5.        Following staining are commonly used in histology lab

    i)                     silver stain

    ii)                    eosin stain

    iii)                 Giemsa stain

    iv)                  Azan stain

    v)                   Hematoxylin stain

    6.        Following statements are true

    i)                     formaldehyde is a gas , 40% of it soluble in water

    ii)                   plastic resins uses as embedding materials

    iii)                 mixture of alcohol and water from 70%-100% causes rehydration

    iv)                  tissue component with negative charge stain more with basic dye

    v)                   Exposure to mild formalin for 8 hrs /per day or 40 hrs in a week is safe.

    7.        Following are the sequence of tissue preparation

    i)                     embedding, fixation , clearing , infiltration

    ii)                   embedding, infiltration, clearing , fixation

    iii)                         fixation, dehydration, clearing, infiltration, embedding

    iv)                  fixation, rehydration, clearing, infiltration, embedding

    v)                   sectioning tissue, fixation, dehydration, clearing, infiltration, embedding

    8.        Following structures are stain blue in hematoxyline and eosin stain

    i)                     Nucleus

    ii)                   Cell membrane

    iii)                 Matrix of cartilage

    iv)                  Cytoplasm

    v)                   Collagen fibres

    9.           Following structures are stain pink  in hematoxyline and eosin stain

    i)                     cytoplasm

    ii)                   nucleus

    iii)                 nucleolus

    iv)                  collagen fibres

    v)                   mitochodria

    10.     Before amounting following steps are done

    i)                     clearing

    ii)                   rehydration

    iii)                 dehydration

    iv)                  tissue section covered by DPX

    v)                   tissue section covered by cover slip