Summary of the Histopathologic Process or Tissue Processing (F) Flashcards
Knowledge about histopath starts where?
In the operation theatre
What are the responsibilities of OT staff?
1) Accurate pt identification
2) Orientation of sxs
3) Adequate fixation
What are the responsibilities of junior pathology staff?
1) Appropriate grossing
2) Sampling
What are the responsibilities of histopathology technicians?
1) Adequate processing
2) Appropriate embedding techniques
3) Microtomy
4) Staining
5) Avoiding unacceptable artifacts
What are the responsibilities of expert pathologists?
1) Inspection of controls
2) Immunohistochemical methods
3) Reporting of the slides
After the sx (/ tissue) is collected, it should be fixed via the use of what fixative?
10% formalin
Formalin causes what?
1) Chemical and physical changes
2) Due to the chemical and physical changes present, it hardens and preserve the tissue
3) Protect from degeneration
Provide an ex of special fixative and where it is used
For testicular biopsies: Bouin’s solution is used
What is highly recommended to be done to sxs?
Tagging
True or False
Every institute should evolve tagging mechanisms
True
Provide an ex of application of the importance of tagging
1 silk suture: for superior margin
2: for lateral margins
What should be done if any orientation of larger sxs are unclear (due to tagging)?
These should be immediately communicated w/ clinician and be clarified
True or False
The communication done if there are any orientation (of larger sxs) that are unclear should be documented by the histotech on the request form
False, because the communication done if there are any orientation (of larger sxs) that are unclear should be documented by the patho on the request form
True or False
Large surgical resection sxs should be sliced by the surgeon
False, because large surgical resection sxs should not be sliced by the surgeon
What are the exs of small fragile sxs?
1) Bone marrow
2) Tru-cut liver
3) Kidney
What should be done to small fragile sxs?
These should be wrapped in a gauze envelope
What are the components of a test requisition form?
1) Pt demographics
2) Details of history
3) Anatomic site of biopsy
4) # of containers sent
5) Sx transportation instructions
Sx should be transported at what temp?
Room temp
What should be present when a sx is transported?
A 10% formalin hazard label
When the sx is going to be transported, where should it be placed?
In a well-sealed leak proof container
What is the importance of sx accessioning?
It accurately labels and track sxs to avoid potential misidentification errors
What are the critical elements in sx accessioning?
Pt and sx identification
How can proper identification of sxs be achieved?
By instructing technicians to use at least 2 pt identifiers when receiving sxs
What are the steps where sx identification is maintained?
1) Sx labeling
2) Grossing
3) Block labeling
4) Slide labeling
What are the responsibilities by the pathologist?
The crucial parts of the pathologic examination w/c are:
1) Precise and systematic gross description
2) Dissection and selection of sections for microscopic study
What should be done to bony or cartilaginous tissues?
These are placed in the decalcifying solution
The bony or cartilaginous tissues are placed in the decalcifying solution for how long?
1 - 7 days
In grossing, the tissue should be washed via the use of what?
Distilled H2O
In grossing, how many times is the tissue washed w/ distilled H2O?
3 - 4 times
True or False
The number of bits present / received in grossing should be noted
True
Where are representative sxs of the tissues placed?
In tissue cassettes
What are the equipment used for placing the representative sxs of tissues in the tissue cassettes?
1) Scalpel
2) Fine pointed forceps
Once the representative sxs of tissues are placed in the tissue cassettes, what should be done to the cassettes?
These should be labeled w/ the identification number
What is integrated in grossing to evaluate the margins of resection on microscopy?
Inking of sxs
True or False
The practice of inking the surgical margins, sampling the tumor margins / capsule, its interface w/ the non tumorous tissue must be followed
True
What should be done to all grossing instruments after grossing?
These are rinsed w/ H2O and wiped clean
What is the ideal place for grossing?
Grossing should be performed in a well-ventilated, well-lit grossing stations
True or False
During grossing, exhaust and proper light are switched on
True
During grossing, what should be worn?
1) Proper gloves
2) Mask
3) Apron
During grossing, what should be used to clean the grossing station?
1% sodium hypochlorite
The grossing instruments are put in what?
1% sodium hypochlorite
True or False
A dedicated grossing room is optional
False, because a dedicated grossing room is mandatory
During grossing (specifically in terms of monitoring), what should be done?
Formalin vapor monitoring
What is the purpose of formalin vapor monitoring?
To detect excess harmful vapor
What are the characteristics of formalin?
1) It is volatile
2) It is toxic
3) It causes irritation to:
a. Eyes
b. Mucous membranes
c. Skin
4) It is associated w/ increased risk for all cancers
What is the meaning of OSHA?
Occupational Safety and Health Administration
What are the exposure limits to formalin accdg to OSHA?
1) 0.75 ppm (as an 8-hr time-weighted average)
2) 2.0 ppm (for short-term [15-min] exposures)
What is the meaning of ppm?
Parts per million
What is the indication if formaldehyde is detected by smell?
It means that exposure is occurring at a concentration beyond acceptable limits
What are the ways on how to limit exposure to formalin?
1) Cover all sx buckets
2) Periodically spray formalin-neutralizing agent on the waste
3) Discard bagged formalin-soaked towels and other waste
4) Cut large fixed organs in a fume hood or downdraft table
5) Monitor the lvl of formalin fumes generated in the hood and outside
6) Level in the hood are kept < 1 ppm
What should be the state of the tissue for it to be cut into 5 um sections?
The tissue should be hardened enough
What is done to 5 um sections of the tissue?
It is stained and examined under a microscope
True or False
H2O should not be removed from the tissue
False, because H2O should be removed from the tissue
After the H2O is removed from the tissue, what is done?
The H2O is progressively replaced by wax, to make a tissue block suitable for sectioning
What are the stages of dehydration?
1) 50% alcohol: 90 mins
2) 70% alcohol: 90 mins
3) 80% alcohol: 90 mins
4) 95% alcohol: 90 mins
5) 100% alcohol: 90 mins
6) 100% alcohol: 12 hrs
How is paraffin impregnation done?
The tissue is immersed in molten wax
In paraffin impregnation, the tissue is immersed in molten wax at what temp?
60 DC
What are the pros of using an automated tissue processor?
1) It saves time
2) It produces consistent results
3) It can overcome:
a. Laborious steps in processing
b. Human errors and forgetfulness
What are the details present in the display (of the automated tissue processor)?
1) Processes for start-up and shut down of the processor
2) Details of solution change
What are the causes of malfxns of automatic processors?
1) Dried sxs
2) Burned sxs
3) Sxs that remained in the clearing agent for too long
How can the dried out tissues / badly processed tissues be best improved?
By reprocessing by the formal-glycerol procedure
True or False
Any malfxns present in the automated tissue processor should all be documented
True
What are the components of the documentation of the malfxn (of the automated tissue processor) that happened?
1) Tissues affected
2) Resource followed
3) Action taken
In an automated tissue processor, what is required to ensure uninterrupted processing?
UPS
What are the results if interruption of carousal movement (of the automated tissue processor) is present?
1) Drying out of tissue
2) Tissue damage
What are the results if shortcuts in processing (via the use of an automated tissue processor) are done?
1) Incompletely dehydrated sxs
2) Cleared and infiltrated sxs
True or False
Mechanical and electrical faults (in automated tissue processors) usually occur
False, because mechanical and electrical faults (in automated tissue processors) occasionally occur
In connection to mechanical and electrical faults (in automated tissue processors), processing mishaps are caused by what?
Human error
True or False
Tissues that are kept for a longer or shorter period than necessary are inadequately processed
True
What is the std working temp for wax?
65 DC
The heating station will automatically switch on at what temp?
< 50 DC
What should be done when working w/ paraffin (specifically at a melting point of < 58 DC)?
Instrument working temp is readjusted
True or False
Temp of the wax bath is checked and recorded monthly
False, because the temp of the wax bath is checked and recorded daily
The use of an inappropriate processing schedule or the making of a fundamental mistake (such as replenishing or sequencing of processing rgnts) can result in what?
It can result in the production of tissue sxs that cannot be sectioned and therefore will not provide any useful microscopic info
The rgnt is changed depending on what?
1) Quality of the sx
2) Quantity of the sx
W/ the load of 300 - 350 cassettes per day, what are the frequencies for changing the ff solutions:
1) Absolute alcohol
2) Xylene
3) Wax
4) Formalin
1) Alternate day
2) Every 3rd day
3) Weekly
4) Everyday
What should be done to the log?
It should be maintained
Why should the log be maintained?
For the days of changing the processing solutions
True or False
The automated tissue processor should be cleaned on a monthly basis
False, because the automated tissue processor should be cleaned on a daily basis
What should be done to the automated tissue processor for once a month?
1) Lift the carousel cover
2) Clean the axle
3) Apply a thin coat of equipment oil
What is anhydrous CuSO4?
Anhydrous copper sulfate
What should be done to anhydrous CuSO4 (in an automated tissue processor)?
1) It should be placed at the bottom of a dehydrating bottle
2) It should be covered w/ 2 - 3 filter papers
What is the action of anhydrous CuSO4 (in an automated tissue processor)?
It removes H2O from alcohol
What is the characteristic of anhydrous CuSO4 (in an automated tissue processor)?
It is white in color
True or False
CuSO4 should be in final alcohol (in an automated tissue processor)
True
What is the characteristic of hydrated CuSO4 (in an automated tissue processor)?
It is blue in color
True or False
Calibrations of the timer and thermostat is required every month
False, because the calibrations of the timer and thermostat is required every yr
In the step of trimming, sectioning, and labeling, what should be done before sectioning?
Wax blocks are rubbed w/ ice
True or False
It is impt to have a properly fixed and embedded block
True
What are the common artefacts that may occur?
1) Tearing
2) Ripping
3) Venetian blinds
4) Holes
5) Folding
True or False
In staining, each procedure that comprises this step is standardized as per requirement
True
True or False
Staining involves multiple steps
True
Since staining involves multiple steps, what should be used?
Work-bench aids
What is done in deparrafinization?
The paraffin is removed by dipping the slides in xylene
In deparrafinization, how many changes of xylene are used and for what time duration is the xylene changed?
3 changes of xylene: used for 5 mins
How is hydration (as a step in staining) done?
The slides are dipped in alcohol
What are the 2 phases where slides are dipped in alcohol (in hydration)?
1st: in 90% alcohol
2nd: in 80% alcohol (for 2 secs each)
After hydration, what should be done to the slides?
Place the slides under running tap H2O for 2 mins
Why should the slides be placed under running tap H2O for 2 mins (after hydration)?
For blueing the hydration of the slide
What should be done to the slides after being placed under running tap H2O?
Stain the slide w/ hematoxylin stain (for 15 - 20 mins)
What should be done to the slide after staining it w/ hematoxylin stain?
Place again the slide under running tap H2O (for 5 mins)
What should be done to the slide after placing it again under running tap H2O?
Dip the slides for a few times in 1% acid-alcohol for decolorization
What should be done after dipping the slides in a decolorizer?
Place the slides under running tap H2O (for 15 mins)
What should be done to the slides after placing these under running tap H2O?
Counterstain w/ 1% eosin stain
What should be done to the slide after counterstaining it?
Immerse the slide in H2O (for 1 min)
What should be done to the slide after immersing it in H2O?
Immerse the slides in ascending concentrations of alcohol (dehydration), then, dip the slides 2 - 3 times (1st: in 90% alcohol; 2nd: in 100% alcohol)
What should be done to the slide after dehydrating it?
Slides are then cleared in xylol thrice (for 2 mins each)
What should be done to the slide after clearing it in xylol?
Mount the slide w/ DPX
How is mounting the slide w/ DPX done?
1) Place some amt of DPX on the tissue section
2) Place a cover slip on it
3) Remove air bubbles by gently pressing w/ a forceps
4) Wipe the excess DPX w/ a tissue
The nuclei of the tissue will stain what color?
Blue
The cytoplasm of the tissue will stain what color?
Shades of pink to red
What is the summary of the steps of the process present in the histopath lab?
1) Sx collection
a. Use of fixatives
b. Use of special fixatives
c. Tagging of sxs
d. Handling of small fragile sxs
e. Presence of test requisition form
2) Sx transportation
a. Should be transported at room temp
b. A 10% formalin hazard label should be present
c. Sxs should be placed in a well-sealed leak proof container
3) Sx accessioning
a. For accurately labeling and tracking sxs
b. For proper identification of sxs
4) Grossing
a. Inking of sxs
b. Gross in a proper room
c. Use proper PPEs when grossing
d. Formalin vapor monitoring (monitoring of environmental conditions)
e. Cleaning of the grossing station
f. Cleaning of the grossing equipment
6) Tissue processing
a. Fix the tissue
b. Since the tissue is already hardened, cut it into 5 um sections
c. These sections are stained and examined microscopically
d. Remove H2O from the tissue (dehydration)
e. Progressively replace the H2O w/ wax (paraffin impregnation)
f. Using automated tissue processor is more beneficial and convenient
g. Certain calibrations must be done
h. Trimming, sectioning, and labeling
A. Honing
B. Stropping
i. Slide preparation
A. Staining
-> Deparaffinization
-> Hydration
-> Dehydration
-> Mount the slide w/ DPX
