Medical Applications of Tissues Flashcards
7 steps of tissue preparation
- Fixation
- Dehydration
- Clearing
- Embedding
- Sectioning
- Mounting
- Staining
Fixation
Formalin Glutaraldehyde
Function–Preservation of normal tissue architecture after death. Fixatives coagulate or precipitate proteins. This renders the structures resistant to further processing.
Dehydration
Chemical used-50-100% of ethanol
Purpose of step-to remove the H2O for the tissue specimen and replace it with alcohol
Clearing
Xylene
Prepares the tissue for the embedding medium
Embedding
Paraffin
Tissus is infiltrated with paraffin so that it is hard enough to section
Sectioning
Tissue is cut into thin sections with a microtome
Mounting
Cut paraffin section is placed on a glass microscope slide
Staining
Various chemicals used
To impact contrast to tissue structures so that they may be distinguished upon microscopic observation
Acid dyes
Chemicals with a net negative charge
Basic dyes
chemicals with net positive charge
Acidophilic
cellular or tissue structures that possess a net positive charge and hence have an affinity for the negative acid dye.
Basophilic
– cellular or tissue structures that possess a net negative charge and hence have an affinity for the positive basic dye.
Metachromasia
a phenonmenon in which a given stain imparts different colors to the tissue
Hematoxylin and eosin (H & E)
Structure Affinity a.H – RNA, DNA, ribosomes, and rER b. E – secretory vesicles, sER, lysosomes, mitochondria, and type I collagen Colors imparted a. H – blue b. E – pink -used for coagulative necrosis
Feulgen reaction
Structure Affinity a. DNA Color imparted a. Magenta (purplish-pink) --can be used to demonstrate nuclear changes in cancer
Mallory triple
Strcture affinity a. Nuclei, muscle, collagen, hyaline cartilage Color imparted a. Nuclei – red b. Muscle – red to orange c. Collagen – blue d. Hyaline cartilage – blue --can be used to highlight fibrosis
PAS reaction
Carbohydrates
Magenta
–can be used in to thickened basement membranes in kidney disease, glycogen storage diseases and in alpha1-antitrypsin deficiency in liver cells (hepatocytes)
Osmic acid
Lipids
Black
Verhoeff
Elastic fibers
Black
–elastic fibers (Marfan syndrome)
Silver methods
Intermediate filaments of nerve cells, glial cells, and reticular fibers
Black
Trypan blue
Macrophages
Blue
Prussian Blue
Hemosiderin (ferric iron)
Blue
–used to demonstrate excessive iron accumulation in hemochromatosis
Nissl
Ribosomes
Blue
Iron hematoxylin
Nuclear elements, chromosomes, mitochondria, centrioles, and muscle striation
Dark blue to black
Congo red
to identify extracellular deposits of amyloid
Immunocytochemistry
A. Technique that uses an antibody tagged with a marker (e.g., fluorescent molecule) directed to a specific antigen (e.g., a protein associated with the cell or tissue).
i. Direct method is less sensitive as fewer antibodies bind to the antigens
ii. Indirect method is more sensitive as more antibodies bind thus amplifying the signal.
Direct method of immunocytochemistry
less sensitive as fewer antibodies bind to the antigens
Indirect method of immunocytochemistry
is more sensitive as more antibodies bind thus amplifying the signal.
Clinical utility of immunocytochemistry
i. Categorization of tumor origin
a. Cytokeratins and epithelial cells
b. Kaposi sarcoma – D2-40 and lymphatic endothelium
ii. Breast cancer
a. Estrogen receptors
b. HER2/neu (aka ERB B2) is a member of the epidermal growth factor receptor family and is overexpressed in some types of breast cancers as well as some adenocarcinomas
c. Carcinoembryonic antigen in breast and intestinal cancers
In situ hybridization
A. Instead of antibodies, this technique uses complementary nucleic acid probes to specifically identify a nuclei acid sequence of interest
B. Can be used to identify a cell infected with a virus (e.g., human papillomavirus type 2) or specific genes on a chromosome (e.g., BRCA1 or BRCA2 genes). The human papillomavirus is a causative agent in cervical and anal cancers and genital warts (e.g., condyloma).
C. Of clinical utility to detect amplification of genes (e.g., Myc genes, HER2/neu). Gene amplification may produce multiple extrachromosomally copies of double minute chromosomes or by another mechanism, amplified genes are interpolated into a region of a chromosome.
