Successfully reported this slideshow.
We use your LinkedIn profile and activity data to personalize ads and to show you more relevant ads. You can change your ad preferences anytime.
PREPARATION OF
HISTOLOGICAL SPECIMENS
• Histology is the study of tissues and how these
tissues are arranged into organs
• Histo in Greek means tissue or web
• ...
TISSUE PREPARATION
• The most widely used method of studying
tissues is using histological slides.
• The tissue in the sli...
TISSUE FIXATION
Aim of Fixation:
1- To prevent autolysis and bacterial attack.
2- To fix the tissues so they will not chan...
Types of Fixative
• Acetic acid
• Formaldehyde 10%
• Ethanol
• Glutaraldehyde
• Methanol
• Picric acid
• Osmic acid (Osmiu...
TISSUE PROCESSING
• Aim:
Is to embed the tissue in a solid medium firm
enough to support the tissue and give it
sufficient...
Dehydration
• To remove fixative and water from the tissue
and replace them with dehydrating fluid.
• Delicate specimens a...
Types of dehydrating agents
• Ethanol
• Methanol
• Acetone
• Tissues may be held and stored indefinitely
in 70% ethanol wi...
Clearing
• Replacing the dehydrating fluid with a fluid
that is totally miscible with both the
dehydrating fluid and the e...
Types of Clearing Agents
• Xylene.
• Toluene.
• Chloroform.
• Benzene.
• Propylene oxide
Embedding
• Is the process by which tissues are
surrounded by a medium such as agar,
gelatin, or wax which when solidified...
Embedding Moulds
A. paper boat mould
B. metal boat mould
C. Dimmock embedding
mould
D. Peel-a-way disposable
mould
E. Base...
CUTTING
• using the microtome
• A microtome is a mechanical instrument
used to cut biological specimens into very
thin sections for microscopic examinat...
Microtome knives
• STEEL KNIVES
• NON-CORROSIVE KNIVES FOR CRYOSTATS
• DISPOSABLE METAL BLADES
• GLASS KNIVES
• DIAMOND KN...
STAINING
Hematoxylin and Eosin (H & E)
• H & E is a charge-based, general purpose stain.
• Hematoxylin stains acidic molecules shad...
Staining Procedure
• Deparaffinize and hydrate to water
• If sections are Zenker-fixed, remove the mercuric chloride
cryst...
Staining machine
• Basic dyes stain:
Heterochromatin
Nucleic acids
Ribosomes
Cartilage
• Acidic dyes stain:
Filaments
Mitochondria
Collagen...
Additional Dyes
• Many tissue components can not be stained
with (Hematoxylene and Eosin).
• Other dyes are used to specif...
Preparation of histological slide
Preparation of histological slide
You’ve finished this document.
Download and read it offline.
Upcoming SlideShare
Tissue processing 2012
Next
Upcoming SlideShare
Tissue processing 2012
Next
Download to read offline and view in fullscreen.

Share

Preparation of histological slide

Download to read offline

Histology

Related Books

Free with a 30 day trial from Scribd

See all

Related Audiobooks

Free with a 30 day trial from Scribd

See all

Preparation of histological slide

  1. 1. PREPARATION OF HISTOLOGICAL SPECIMENS
  2. 2. • Histology is the study of tissues and how these tissues are arranged into organs • Histo in Greek means tissue or web • Tissues consist of cells and extra cellular matrix • The function of the tissue depends on the interaction between cells and extracellular matrix. • The small size of cells and matrix content make the study of tissues dependent on microscope and other advances in biological techniques
  3. 3. TISSUE PREPARATION • The most widely used method of studying tissues is using histological slides. • The tissue in the slide must reflect the actual nature of the tissue in the body • To insure that, tissues to be studied must pass through a series of steps before examination • These steps are in sequential order • Fixation is a complex series of chemical events that differ for the different groups of substance found in tissues.
  4. 4. TISSUE FIXATION Aim of Fixation: 1- To prevent autolysis and bacterial attack. 2- To fix the tissues so they will not change their volume and shape during processing. 3- To prepare tissue and leave it in a condition which allow clear staining of sections. 4- To leave tissue as close as their living state as possible, and no small molecules should be lost. •Fixation is a reaction between the fixative and proteins in the specimen which form a gel, so keeping every thing as their in vivo relation to each
  5. 5. Types of Fixative • Acetic acid • Formaldehyde 10% • Ethanol • Glutaraldehyde • Methanol • Picric acid • Osmic acid (Osmium tetroxide)
  6. 6. TISSUE PROCESSING • Aim: Is to embed the tissue in a solid medium firm enough to support the tissue and give it sufficient rigidity to enable thin sections to be cut , and yet soft enough not to damage the knife or tissue. • Stages of processing: 1.Dehydration. 2- Clearing. 3- Embedding.
  7. 7. Dehydration • To remove fixative and water from the tissue and replace them with dehydrating fluid. • Delicate specimens are dehydrated in a graded ethanol series from water through 10%-20%-50%-95%-100% ethanol to minimize tissue distortion from diffusion currents. • In the paraffin method, dehydration from aqueous fixatives is usually initiated in 60%- 70% ethanol, progressing through 90%-95% ethanol, absolute ethanol before proceeding to the clearing stage.
  8. 8. Types of dehydrating agents • Ethanol • Methanol • Acetone • Tissues may be held and stored indefinitely in 70% ethanol without harm
  9. 9. Clearing • Replacing the dehydrating fluid with a fluid that is totally miscible with both the dehydrating fluid and the embedding medium. • Choice of a clearing agent depends upon many factors
  10. 10. Types of Clearing Agents • Xylene. • Toluene. • Chloroform. • Benzene. • Propylene oxide
  11. 11. Embedding • Is the process by which tissues are surrounded by a medium such as agar, gelatin, or wax which when solidified will provide sufficient external support during sectioning. • A substances added alone or in combination to the wax to: Improve ribboning. Increase hardness. Decrease melting point Improve adhesion between specimen and wax
  12. 12. Embedding Moulds A. paper boat mould B. metal boat mould C. Dimmock embedding mould D. Peel-a-way disposable mould E. Base mould used with embedding ring ( F) or cassette bases (G)
  13. 13. CUTTING • using the microtome
  14. 14. • A microtome is a mechanical instrument used to cut biological specimens into very thin sections for microscopic examination. • Most microtomes use a steel blade and are used to prepare sections of animal or plant tissues for histology.
  15. 15. Microtome knives • STEEL KNIVES • NON-CORROSIVE KNIVES FOR CRYOSTATS • DISPOSABLE METAL BLADES • GLASS KNIVES • DIAMOND KNIVES
  16. 16. STAINING
  17. 17. Hematoxylin and Eosin (H & E) • H & E is a charge-based, general purpose stain. • Hematoxylin stains acidic molecules shades of blue. • Eosin stains basic materials shades of red, pink and orange. • H & E stains are universally used for routine histological examination of tissue sections.
  18. 18. Staining Procedure • Deparaffinize and hydrate to water • If sections are Zenker-fixed, remove the mercuric chloride crystals with iodine and clear with sodium thiosulphate (hypo) • Mayer's hematoxylin for 15 minutes • Counterstain with eosin from 15 seconds to 2 minutes depending on the age of the eosin, and the depth of the counterstain desired • Dehydrate in 95% and absolute alcohols, two changes of 2 minutes each or until excess eosin is removed • Clear in xylene, two changes of 2 minutes each • Mount in Permount or Histoclad
  19. 19. Staining machine
  20. 20. • Basic dyes stain: Heterochromatin Nucleic acids Ribosomes Cartilage • Acidic dyes stain: Filaments Mitochondria Collagen Muscle fibers
  21. 21. Additional Dyes • Many tissue components can not be stained with (Hematoxylene and Eosin). • Other dyes are used to specifically stain certain tissue components Resorcin-Fuchsin for elastic fibers Silver stain for reticular fibers and basement membrane Periodic-Acid Schiff (PAS) Reaction for CHO
  • haivishwa

    Oct. 11, 2021
  • BravoIronSheet

    Sep. 30, 2021
  • AmirHussain126

    Jul. 10, 2021
  • DentMaRyem

    Jun. 26, 2021
  • BethwelRuto

    May. 20, 2021
  • EmanAbdullah33

    Mar. 4, 2021
  • TruptiJadhav24

    Jan. 23, 2021
  • SriDharani7

    Jun. 7, 2020
  • GxadxerBaxir

    May. 13, 2020
  • PriyapooraniRanga

    May. 2, 2020
  • muhsinalghezzi

    Mar. 4, 2020
  • YassirBassim

    Feb. 15, 2020
  • hemantkumar307

    Feb. 9, 2020
  • SmitaChaudhari2

    Oct. 14, 2019
  • NgmaapuoriDouglas

    Oct. 8, 2019
  • SheetalPunia1

    Sep. 9, 2019
  • KrishnamRagaPavitra

    Jun. 26, 2019
  • SaraEsamMousa

    Jun. 14, 2019
  • Shiwaangisharma

    May. 17, 2019
  • MennaJameel

    Apr. 12, 2019

Histology

Views

Total views

24,303

On Slideshare

0

From embeds

0

Number of embeds

748

Actions

Downloads

694

Shares

0

Comments

0

Likes

53

×