Tissue Prep And Staining Flashcards
What is the most common fixative and what is it good for?
Formalin and it’s good for viewing larger objects not fine cytological detail.
What are acid fixatives used to view?
Chromatin, nucleoli, and spindle fibers
Name 3 acid fixatives
Carnoy’s fluid
Zenker’s fluid
Bouins’s fluid
What are basic fixatives used for? And name 1 type
Mitochondria staining
Zirckle Erliki
What comes after fixing and what is it’s purpose?
Dehydrating, placing the tissue increasing strengths of ethanol. The purpose is to remove all water from the specimen because it will be embedded in paraffin (hydrophobic)
What shouldn’t you use ethanol for dehydration when your goal is to observe fats?
Because ethanol dissolves fats.
What is the purpose of clearing? And how do you accomplish clearing?
The goal is to remove the alcohol and you do so by replacing it with xylene or cedar oil.
What are the fixatives used for TEM?
Glutaraldehyde and osmium tetroxide
Describe the process of embedding
The specimen gets 3 paraffin baths
Placed in paraffin mold
Hardened by a cold water bath
Describe the embedding steps for TEM
Tissues are infiltrated with monomeric resin
Resin is polymerized
Tissue sample are less them 1mm cubed.
How do you prepare thin slices of specimen
Rotary microtome
How do you d sectioning for TEM
Sections are 50 to 150 nm
Sections so fragile must be floated onto plastic coated copper mesh grid
Holes in copper grid allow for beam of electrons to pass through tissue sample.
Diamond knives are used
How do you prep for staining?
Remove paraffin with xylene Remove xylene Stains are applied Dehydration Removal of alcohol with xylene Cement and cover slip
What is the H & E stain useful for?
Displaying structures, not chemical characteristics. H acts as a basic dye and stains light blue or purple and E acts as an acid dye and stains yellowish or pinkish color.
What are basic dyes
They bind with basic parts of the tissue (phosphate, sulfate and carboxyl groups).