1/6/2020
The Tissue Level of Organization
• Group of similar cells
– common function
• Histology
– study of tissues
• Pathologist
– looks for tissue changes
that indicate disease
Four Basic Tissues
• Epithelial Tissue
– covers surfaces because cells are in contact
– lines hollow organs, cavities and ducts
– forms glands when cells sink under the surface
• Connective Tissue
– material found between cells
– supports and binds structures together
– stores energy as fat
– provides immunity to disease
1
1/6/2020
Four Basic Tissues
• Muscle Tissue
– cells shorten in length producing movement
• Nerve Tissue
– cells that conduct electrical signals
– detects changes inside and outside the body
– responds with nerve impulses
Epithelial Tissue -- General Features
• Closely packed cells forming continuous sheets
• Cells sit on basement membrane
• Apical (upper) free surface
• Avascular---without blood vessels
– nutrients diffuse in from underlying connective
tissue
• Rapid cell division
• Covering / lining versus glandular types
2
1/6/2020
Basement Membrane
• holds cells to connective
tissue
Types of Epithelium
• Covering and lining epithelium
– epidermis of skin
– lining of blood vessels and ducts
– lining respiratory, reproductive, urinary & GI
tract
• Glandular epithelium
– secreting portion of glands
– thyroid, adrenal, and sweat glands
3
1/6/2020
Connective Tissues
• Cells rarely touch due to extracellular matrix
• Matrix(fibers & ground substance secreted by cells)
• Consistency varies from liquid, gel to solid
• Does not occur on free surface
• Good nerve & blood supply except cartilage & tendons
Mature Connective Tissue
• Loose connective tissue
• Dense connective tissue
• Cartilage
• Bone
• Blood
• Lymph
4
1/6/2020
Muscle
• Cells that shorten
• Provide us with motion, posture and heat
• Types of muscle
– skeletal muscle
– cardiac muscle
– smooth muscle
Nerve Tissue
• Cell types -- nerve cells and neuroglial (supporting) cells
• Nerve cell structure
– nucleus & long cell processes conduct nerve signals
• dendrite --- signal travels towards the cell body
• axon ---- signal travels away from cell body
5
1/6/2020
6
1/6/2020
The Hematoxylin and Eosin Stain
• Long history of use–Mayer 1904
• Primary diagnostic technique in the histo-
pathology laboratory
• Primary technique for the evaluation of
morphology
• Widely used –2.5 to 3 million slides per day
7
1/6/2020
The Hematoxylin and Eosin Stain
• The Hematoxylin and Eosin combination is
the most common staining technique used
in histology
• The diagnosis of most malignancies is
based largely on this procedure
• Pathologist expectations or preferences are
extremely subjective
Some Basics of Dye Chemistry
• Why do dyes stain specific elements of cell
and tissues?
• Dyes demonstrate an affinity for molecules
within cells and tissues
• Affinity is the result of attractive forces
between the dye molecule and molecules
within the tissue
– Dyes have a greater affinity for tissue
molecules than solvent molecules
8
1/6/2020
Some Basics of Dye Chemistry
• The affinity of dyes for tissue elements is
affected by a number of factors
– The structure of the dye molecule
– The shape of the dye molecule
– The charge distribution of the dye
– The solvent the dye is dissolved in
Some Basics of Dye Chemistry
• Acidic pH (< 7) ***Acidophilic Stains
– Higher concentration of H+
– Some tissue molecules will bind H+ and will
carry a positive charge
• Basic pH (> 7) ***Basophilic Stains
– Lower concentration of H+
– Some tissue molecules will give up H+ and will
lose a positive charge
9
1/6/2020
Some Basics of Dye Chemistry
• Charge distribution of the dye determines
attractive or repulsive characteristics of the
dye
• Positive charges (+)
– Cations
– Attracted to negatively charged molecules
• Negative charges (-)
– Anions
– Attracted to positively charged molecules
• Charge is largely determined by pH
10
1/6/2020
Hematoxylin
• Natural dye extracted from the wood of the
Logwood tree found in Mexico and Central
America
• Hematoxylin has little or no staining
capacity
• Hematoxylinis oxidized to hematein
Hematoxylin
• Oxidation of hematoxylin to hematein
– Air/light
– Mercuric oxide
– Sodium iodate
• Today almost all hematoxylins are oxidized
by sodium iodate
• Oxidation continues after formulation is
complete
11
1/6/2020
Basic Chemistry
• Hematoxylin solution is a mixture
– Hematoxylin
– Hematein
– Aluminum ions
– Hematein/aluminum
– Solvent mixture
Basic Chemistry
• Hematein bound to aluminum has an overall
positive charge
• Hematein/aluminum binds negatively
charged molecules in cell and tissues
• Chromatic material in cell nucleus is
strongly negative in it’s ionic charge
12
1/6/2020
Basic Chemistry
• Specific staining:
– Nuclei; DNA in
chromatin
• Non specific -
Background
staining:
– Cytoplasm
– Goblet cells of
G.I. tract; mucin
Hematoxylin - Types
• Commercially available hematoxylins:
– Harris – Most Common - Purple
– Mayers – Purple/blue
– Gill I, II, III - Blue
– Weigert’s - Black
– Proprietary formulas
13
1/6/2020
Eosin
• Unlike hematoxylin, eosin is a synthetic
dye.
– Derived from fluorescein
• Eosin Y is a member of the xanthene family
of dyes
– Eosin Y
– Eosin B
– Phloxine B
– Fluorescein
Eosin
• Eosin is acidic
– Negatively charged
• Prepared in alcohol solvent
• Eosin binds to positively charged proteins in
the cytoplasm and in connective tissue
14
1/6/2020
The H and E Stain
• The overall coloration of the stained
specimen is the result of the balance of the
intensity of the alum-hematoxylin and eosin
• Remember!
– Alum-hematoxylin – Nuclei, but also some
cytoplasm
– Eosin Y – Cytoplasm, but also some nuclei
15
1/6/2020
16
1/6/2020
Introduction to Special
Staining
Purpose of Special Staining
• To demonstrate tissue elements or
pathogenic states that are not demonstrated
by the H&E stain
17
1/6/2020
Utilization
What the stains show in the same tissue....
– H&E
– Trichrome
– Silver reactions
– Verhoeff van Gieson
– PAS
– Oil red O
Liver
H&E PAS - glycogen
18
1/6/2020
Liver
H&E Reticulin fiber
Muscle
H&E Trichrome
19
1/6/2020
Intestinal Wall
H&E Trichrome
Kidney Glomerulus
H&E PAS
20
