histology: methods of study

first lecture tissue prep tissue strain artifacts plane of section useful tip

what is histology

study of microscope structure and organization of tissues of the body relative to their function

methodology of histology

1. examine slide, recognize components

2. assess if normal

3. if abnormal, identify what is different

before tissue stain what must you do

tissue prep: collect and fix immediately

paraffin for thin sectioning

then you stain, visualize, and view under electron microscope: transmission or scanning EM

what three light microscopes do not require tissue staining?

dark field (DF)

phase contrast (PC)

differential interference contrast (DIC)

what is the light microscope type that typically requires tissue staining?

bright-field (BF) 95% of what we will see

positive/cationic/basic stains will bind to negative or positive molecules?

negative and stain purple

negative tissues are basophilic and have an affinity for hematoxylin; lots of proteins

negative/anionic/acidic stain will bind to negative or positive molecules?

positive and stain pink

positive tissues are acidophilic and have an affinity for eosin

why would other stains be used?

cytoplasmic and ECM proteins cannot be distinguished with H&E staining

some specific cells/inclusions are not preserved with fixation- appear empty

ex: lipid stains for steatosis (fatty liver disease) to see adipocytes are full of lipids

mucus stains

periodic-acid schiff (PAS) stains molecules rich in carbs (glycoproteins and proteoglycans)

silver stains

bind reticular fibers and nerve cell processes

Masson’s trichrome

three-colored stain that distinguishes cells from surrounding ECM

• blue=collagen (connective tissue)

• purple/dark= nuclei(-)

• cytoplasm/keratin= red

elastic fiber stain

aorta and elastic cartilage

metachromasia

change in color within cell/tissue due to interaction with basic dye and polyanions

basic dye=toluidine blue, for mast cells the nuclei is dark but lighter than the outside which is super negative due to mast cell granules which hold heparin

explain a fluorescent tag

anti-tubulin antibody has a fluorescent tag. it binds to tubulin which is the component of cilia.

ground section

take a cut and grind small enough for light to pass through

no organic tissue; this is mineralized bone

air spaces are dark (previous organic)

fine detail visible

non-mineralized bone viewing

no mineralized tissue

soft flexible tissue stains well with H&E

fine detail less visible

three tissues of the teeth

dentin (70%) and cementum (60%) have mix of collagen and mineralized tissue and resemble bone

enamel (95%) : little protein and no collagen

%=inorganic

tooth composition

pulp (100% organic), periodontal ligament and the three tissues

helpful tips

1. look at entire field of view

2. notice scale (compare to nuclei or RBCs)

3. note nuclear and cytoplasmic characteristics

4. ratio between cell and non-cell

5. diagnose epithelium when present: nuclei, type, homo or hetero, unique features: cilia or microvili