Histology (the basics) Flashcards
What is histology?
- The microscopic study of tissue
- maintains the tissue architecture and can encompass multiple cell types
What are the uses of histology?
- Biopsy - representative sample of tissue
- Thin sectioning - to avoid unclear stack of cells
- Dyes - to allow cell types/proteins to be visible under microscope
What is cytology?
- The study of cells
- tends to focus on single cell type away from tissue (lavage, aspiration, skin scrape)
Histology can tell you about the function of a cell based on what?
- shape
- polarisation
- intracellular contents
- location compared to other cells/structures
- if a cells are failing in a function
What are the stages to produce sections of tissue for light microscopy?
- fixation
- processed
- sectioned
- stained
Without processing tissue for microscopy what would happen?
- it is not permanent so tissue decays
- its difficult to cut thin sections to allow light to pass through
- it lacks colour
Fixative for the preservation of tissue is chosen depending on what?
- the size of the tissue
Describe fixation (1)
- fixed in formon (24hrs)
- stored in 70% alcohol
- original sample = trimmed, transferred into plastic cassette then into a stainless steel carrier
Describe Processing (2)
- to remove water the tissue samples go through increasing strengths of alcohol sol.
- water in tissue is replaced with molten paraffin wax
- tissue dehydrated gradually to avoid shrinkage
- tissue exposed to clearing agent to make it translucent
Describe embedding (part of processing tissue)
- specimen immersed in paraffin wax (1hr)
- tissue removed and put in metal mould filled with molten wax - more wax added on top
- placed on cold plate and solidifies, them removed from mould.
Describe sectioning
- sections cut using a rotary microtome
- tissue clamped into microtome, knife inserted and clamped in holder
- stage is locked and dial set at required thickness
- the sections of wax and tissue form a ribbon that can be lifted off Kinfe and laid down in strips
- sections collected onto slide and dried in oven 40.C
- dewaxing and rehydrating is next to make slides ready to stain
Describe staining
- Haematoxylin and eosin (H&E)
- haematoxylin stains nuclei a blue-black colour
- eosin stains cytoplasm and intercellular components pink
- the blue dye is removed by 1% hydrochloric acid solution until the nuclei can be seen clearly (differentiation)
- then placed in weak ammonium to produce deep blue colour
- slide then stained with eosin, cytoplasm and intercellular contents = red, pink
- once staining is complete it looks purple and pink
Describe how sections are made permanent after staining
- dehydrated in alcohol and cleared in clearing agent to make it translucent
- PIX is a mounting medium and adheres the cover slip to the section
- left in oven to dry overnight
What type of microscope does histology use
- light microscope
what is a longitudinal section?
- A section that is cut along the long axis of a structure
What is a transverse section?
- a section that is cut perpendicular to the longest axis of a structure
What is an oblique section?
- A cut that is at any angle between the longitudinal and transverse planes
What does Haematoxylin dye?
- Basic blue dye
- stains nuclei mainly (nuclei acid) a blue-purple
What does eosin stain?
- acidic dye
- stains cytoplasm mainly (proteins) pink
What other common dye can be used in histology?
- Giemsa Stain
What is Methylene blue?
- A basic dye
- It is positively charged, so binds negatively charged molecules
What dyes are used in blood smears?
- Methylene blue
- Azure B
- Eosin
What is Azure B and eosin?
- acidic dyes
- -ve charge, so binds positively charged molecules
