INTRODUCTION TO HISTOLOGY 7.

Trimming
Histology
- the study of the tissues of the body and
how these tissues are arranged to
constitute organs
PREPARATION OF TISSUES FOR
STUDY
Biopsy: tissue examination from alive
Autopsy: tissue examination from dead
1. Gross Examination
- tissue samples from the organ
- describing the specimen
- placing all or parts of it into a small
plastic tissue cassette which holds the
tissue while it is being processed to a
paraffin block
2. Fixation
- small pieces of tissue are placed in
solutions of chemicals (fixatives)
- cross-link proteins and inactivate
degradative enzymes, which preserves
cells and tissue structure
- most common fixative: formaldehyde
- paraffin block is trimmed to expose the
tissue for sectioning (slicing) on a
microtome
8. Tissues Sectioning
- sectioning (slicing) of tissue using a
microtome
- placed in floatation water bath to
minimally melt the paraffin
9. Staining
- addition of staining dyes to visualize the
tissue structures under the microscope
- most common:
Hematoxylin and Eosin dye (H&E)
- acidophilic and basophilic dyes
10. Mounting
- addition of mounting medium to the
stained tissues and cover with cover slip
to preserve the tissues
- adhesive and preservative

3. Dehydration
- tissue is transferred through a series of
increasingly concentrated alcohol solutions
(dehydrating solutions), ending 100%,
which removes all water
- increasing concentration dapat (from (small intestine under H&E stain)
30%, 50%, 70%, 90%, absolute alcohol)
- most common dehydrator: ethanol MICROSCOPY
- remove water since we are dealing with
water insoluble solutions Two major categories:

4. Clearing A. LIGHT MICROSCOPY

- alcohol is removed in organic solvents
(clearing agents) in which both alcohol
and paraffin are miscible
- remove alcohol
- commonly used: xylene (5 series)
 Bright-Field Microscopy
- stained tissue is examined with
ordinary light passing through the
preparation

5. Infiltration
- tissue is the placed in melted paraffin
(impregnating agent) until it becomes
completely infiltrated with this substance
- using paraffin oven

6. Embedding
- paraffin-infiltrated tissue is placed in a
small mold with melted paraffin and
allowed to harden
- paper boat mold sa pinas bc pobre
 Fluorescence Microscopy  Polarizing Microscopy
- tissue sections are usually irradiated - produces an image only of material
with ultraviolet (UV) light and the having repetitive, periodic
emission is in the visible portion of the macromolecular structure
spectrum - strike a specific pathway
- fluorescent substances (dyes or
lamps) appear bright on a dark
background
- has a source of UV or other light and
filters that select rays of different
wavelengths****

B. ELECTRON MICROSCOPY
- supreme

 Transmission Electron
Microscopy (TEM)
- magnified as much as 400,000 times
to be viewed in detail

 Phase-Contrast Microscopy
- uses a lens system that produces
visible images from transparent
objects
- unstained preparation
- based on the principle that light
changes its speed when passing
through cellular and extracellular
structures*****

 Scanning Electron Microscopy

- produces and focuses a very narrow
 Confocal Microscopy
beam of electrons, but in this
- high resolution and sharp focus than
instrument the beam does not pass
bright-field microscopy
through the specimen
- computer aided microscope
- the surface of the specimen is first
- eliminate stray and scattered light
dried and spray-coated with a very
using confocal pinholes and splitter
thin layer of heavy metal (often gold)
which reflects electrons in a beam
scanning the specimen
- reflected electrons are captured by a
detector, producing signals that are
processed to produce b&w images
- computer aided