H&E Flashcards
highlight and differentiate tissue components and allow them to be seen under the microscope
biological dyes
most common staining technique used in histology
hematoxylin and eosin
dyes have a greater affinity for _______ molecules than solvent molecules
tissue
the result of attractive forces between the dye molecule and molecules within the tissue
affinity
affinity of dyes for tissue elements is affected by 4 factors
-structure of dye molecule
-shape of dye molecule
-charge distribution of dye
-solvent characteristics
determines attractive or repulsive characteristics of the dye
charge distribution
attracted to negatively charged molecules
hematoxylin
attracted to positively charged molecules
eosin
pH largely determines this
charge
pH of hematoxylin should be
2.2-2.8
oxidation of hematoxylin forming the active dye ingredient
hematein
hematoxylin is a natural dye extracted from the wood of this tree
Logwood
this can oxidize hematoxylin to hematein
sodium iodate
oxidation can also occur with this exposure
air
hematein produces a precipitate that must be _________ before use
filtered
this is required to help link hematoxylin to a desired tissue
mordants
metal salts often used in histology as a mordant
aluminum and iron
need hematoxylin and hematein mixture as this occurs
oxidation
need ______ to dissolve the dry powder dye which acts as a carrier of the dye
solvent such as water
example of progressive staining
frozen section
tissue is left in the staining solution just long enough to reach desired endpoint
progressive staining
deliberately overstaining where the dye completely saturates all tissue elements
regressive staining
a popular regressive stain
Harris Hematoxylin
this is preferred when a very clear differentiation of tissue elements is desired
regressive staining
tissue is selectively destained using this process
differentiation
you need to do this in progressive staining
monitoring, to see if it is staining correctly
this is achieved by using a dilute acid/acid alcohol
differentiation
removes excess background staining
differentiation
the ions in the differentiation solution diffuse more rapidly than the dye molecule which causes what
release of the loosely attached dye
differentiation is halted by this when the end point is reached
water rinse
the process of shifting the color from purplish to blue by the application of a weak alkaline solution
bluing
bluing agents are typically _______ with a pH of 7.5-9.0
alkaline
enhance the contrast of H&E by increasing the crispness of hematoxylin
bluing agents
used in both progressive and regressive staning
bluing
counter stain used to demonstrate the general architecture of the tissue and provide contrast to the now stained nuclei
eosin
acidic dye that binds to the basic parts of the cell (cytoplasm)
eosin
erythrocytes and keratin will be this color with eosin
red or orange
collagen will be this color with eosin
pink
smooth muscle will be this color with eosin
dull pink
pH of eosin
4.0-4.5
eosin should have how many shades under a scope
3
what should hematoxylin depict under a scope
nuclear membrane and stained chromatin
rushing through this step can result in spotty staining
deparaffinization
too light hematoxylin occurs due to (4)
-too many slides
-short staining time
-overdecalcified removing nucleic acids
-autolysis or poor fixation
too dark hematoxylin occurs due to (4)
-long staining time
-short destaining time
-laser surgery
-metallic sheen from overoxidation
too light eosin can occur if (2)
-alcohol rinse after stain was too long or too aqueous
-short staining time
one tone of eosin indicates (2)
poor fixation or overstaining
