LEC6 - INTRO TO HISTOPATHOLOGIC TECHNIQUES Flashcards
is defined as the process of preparing the tissue
tissue processing
refers to a better and more effective means of studying tissues whether normal or abnormal is by examination of their sections and smears which have been permanently preserved, stained, and mounted on glass slides with cover slips for permanent keeping
- Processing of Tissue
- Why examine histopathologic specimens?
- Pathology attempts to explain the WHYs and wherefores of the signs and symptoms manifested by patients while providing a sound foundation for rational clinical care and therapy.
- Determine if the sample is benign or malignant
is the microscopic study of the normal tissues of the body
histology
is the microscopic study of tissues affected by disease
histopathology
procedures adopted for the preparation of material for such studies
histologic or histopathologic technique
the examination of tissue has 2 major type of tissue
fresh and preserved
tissue processing is a process, by means of process, it refers to the phases which are
pre analytic
analytic
post analytic
the method of tissue examination may vary according to the following
the structural and chemical components of the cells to be studied
the nature and amount of the tissue to be evaluated
2 types of tissue to be examined
fresh and frozen tissue
- ___ are usually examined when there is an immediate need for evaluation
Fresh Tissues
- Advantages of fresh tissues
o Examined in the living state thereby allowing protoplasmic activities such as: motion, mitosis, phagocytosis and pinocytosis to be observed
again, what are the protoplasmic activities we can observed if we are using fresh tissues
motion, mitosis, phagocytosis, and pinocytosis
- Disadvantages of the use of fresh tissues
its use has been limited/ the protoplasmic activities are limited in time only
liable to develop the changes that have been usually been observed after death
advantage of preserve tissue
once the cellular activity of tissue has stopped is preserved in life-like manner
METHODS OF FRESH TISSUE EXAMINATION
● Teasing or Dissociation
● Squash Preparation (Crushing)
● Smear Preparation
○ Streaking
○ Spreading
○ Pull-apart
○ Touch preparation (Impression smear)
● Frozen Section
A method in fresh tissue examination whereby a selected tissue specimen is immersed in a watch glass containing isotonic salt solution
Teasing or Dissociation
hypertonic solution can cause the cell to
shrink
hypotonic solution can cause the cell to
swell
teasing or dissociation method of fresh tissue is examined using what microscopy
unstained and examined using phase contrast and bright field microscope
if we want to stain the specimen by examining it using teasing or dissociation method, we can use ____ dye
methylene blue dye
CRUSHING method is also called as
squashing preparation
a process whereby a small piece of tissue in is placed in microscopic slide and forcible compressed with another slide or with a cover glass
squash preparation (crushing)
how many diameter is the limit for squash preparation
not more than 1 mm
can we use stain in squashing preparation? if so, how do we stain it
a vital stain may be placed at the junction of the slide and the cover glass, and allowed it to get absorbed by the tissue through capillary attraction
- The process of examining sections or sediments whereby cellular materials are spread lightly over a slide by means of a wire loop or applicator, or by making an apposition smear with another slide
smear preparation for fresh tissue examination
this type of fresh tissue method of examination is useful in cytologic examination particularly for cancer diagnosis
smear preparation
techniques in smear preparation
streaking
spreading
pull apart
touch preparation
this smear preparation technique that uses an applicator stick or platinum loop, the material is rapidly and gently applied in a direct or zigzag line throughout the slide
streaking
what should we avoid in streaking - smear preparation
making of too thin or too thick smears, since they make the tissues unsuitable for examination
streaking -smear prep is used for preparing
mucoid secretions
vaginal secretions
sputum
gastric content
streaking - smear prep uses what materials
spatula,
dissecting needle
applicator stick
this smear prep attempts to obtain a relatively uniform distribution of secretion
streaking
smear prep technique
o A selected portion of the material is transferred to a clean slide and gently spread into a moderately thick film by teasing the mucous strands apart with an applicator stick
spreading
a smear prep that is A little more tedious but maintains cellular interrelationships of the material to be examined
Spreading
the use of spreading technique in smear prep
for fresh sputum
bronchial aspirates
thick mucoid secretions
o A special method of smear prep whereby the surface of a freshly cut piece of tissue is brought into contact and pressed on to the surface of a clean glass slide,
Touch preparation (Impression smear)
touch preparation is also called as
impression smear
a smear prep which it allows the cells to be transferred directly to the slide for examination
touch preparation or impression smear
Touch preparation (Impression smear) is examined using
phase microscope or stained for light microscope
o Cells may be examined without destroying their actual intercellular relationship
Touch preparation (Impression smear)
smear prep technique used in csf
pull apart
smear prep technique used in breast secretion
pull apart
method of rapid tissue processing
frozen section
this type of fresh tissue prep recommended for lipids and nervous tissue
frozen section
thickness of frozen section
10-15 um
a cold chamber kept at an asmospheric temperature of -10C to -20C
cryostat
- Frozen sections are commonly used for:
o Rapid pathologic diagnosis during surgery
o Diagnostic and research enzyme histochemistry
o Diagnostic and research demonstration of soluble such as lipids and carbohydrates
o Immunofluorescent and immunohistochemical staining
o Some specialized silver stains, particularly in neuropathology.
3 section of histopath
paraffin section
frozen section
Celloidin section
- Commonly used methods of freezing
o Liquid nitrogen
o Isopentane cooled by liquid nitrogen
o Carbon dioxide gas
o Aerosol sprays
advantage of liquid nitrogen
used in histochemistry
most rapid of the commonly available freezing agents
disadvantage of liquid nitrogen
soft tissue is liable to crack producing ice crystals or freeze artifacts
causes vapor phase
advantage of isopentane cooled by liquid nitrogen
excellent method for freezing muscle tissue
advantage of carbon dioxide gas
adapting a conventional freezing microtome
advantage of aerosol sprays
adequate for freezing small pieces
rapidly freezing blocks of any type of tissue
2 method of preparing frozen section
cold knife procedure
cryostat procedure
a method of preparing frozen section wherein a tissue blocks are frozen by adapting a conventional freezing microtome gas supply of carbon dioxide gas from a CO2 cylinder or by using a specially made piece of equipment known as cryostat
cold knife procedure q
optimum working temperature of cryostat is
-18C to -20C
cryostat consist of
insulated microtome housed in an electrically driven refrigerated chamber and maintained at a temperature near -20*C where microtone, knife, specimen and atmosphere are kept at the same temperature
steps in processing for paraffin section
fixation
decalcification(optional)
dehydration
clearing
infiltration (impregnation)
embedding
trimming
section-cutting
staining
mounting
labelling
conventional method for freezing section
carbon dioxide gas
clearing is also called as
dealcoholization
Criteria for rejection of gross specimen
- Discrepancies between the requisition and specimen label
- Specimen with no labels, or mislabeled
- Leaking specimen container
- Absent clinical data or history
- Inappropriately identified specimen
can Specimens be received fresh (without fixative) or in formalin
yes
For frozen sections, tissue is received _____ for immediate microscopic evaluation by the Pathologist.
fresh
. May be received for an OR consultation
fresh tissue specimen
- The Pathologist will look at the specimen and make a gross diagnosis
.what are the roles of PA
c. As the PA, you may weigh and measure the specimen.
d. As the PA, you may also ink the specimen, if necessary; however,
always check with the Pathologist before making any of these decisions.
Ideally, the specimen should have at least _____times x its volume of formalin
20
Specimen Accessioning rules
A. Each specimen receives an accession number.
B. Each number is unique to that particular case and is NEVER reused.
C. The specimen container(s), the requisition slip and all cassettes are labeled with the case/accession number
Grossing, often referred to as “______”, it involves a careful examination and description of the specimen
cut-up
grossing includes careful examination and description of the specimen such as it’s _______
appearance,
number of pieces
dimensions
- the most important processes in which the pathologist arrives at a diagnosis.
Gross Examination of Specimens
- Steps in grossing
- Identification of the specimen
- Patient’s surname, name birthday, hosp. number
- The spec container must bear the same name and acc. Number in the request form
Responsibility of a technician
- Specimen preservation.
- Specimen labeling, logging and identification.
- Preparation of the specimen to facilitate their gross and microscopy.
- Record keeping
- orientation markers
INKS
NICKING
SUTURE ATTACHES (LL and SS)
an orientation marker that is to identify and orient the spec component
inks
orientation marker for indicating laterality
nicking
suture attaches are represented as
LL and SS
long lateral
short superior
The specimen is then cut into representative sections and is put in small plastic cassette to hold the tissue
size of cassette
- 3.0x2.5x0.4 cm
- Spec should not be more than _______ mm in thickness.
0.3
Specimen-types
excision-specimens
incisional biopsies
punch biopsies
shave biopsies
curettings
core biopsies
where whole organs or affected areas are removed at operation
- Excision specimens (surgical biopsies
, where tissue is removed for diagnosis from within an affected area
Incisional biopsy specimens,
, to remove a small piece of suspicious tissue for examination (often from the skin)
Punch biopsies
where small fragments of tissue are “shaved” from a
surface (usually skin
- Shave biopsies
where tissue is removed in small pieces from the lining of the uterus or cervix
- Curettings
where a small tissue sample is removed using a special needle sometimes through the skin (percutaneously
- Core biopsies
