Quality Control in

Histopathology
 The quality control checks in a histopathology lab will include
accurate patient identification, fixation, adequate processing,
appropriate embedding techniques, microtomy, unacceptable
artifacts and inspection of controls to determine correctness of
special stains and immunohistochemical methods.
 . It is the responsibility of the pathologist to perform the final
quality control examination as they read the slide and determine
whether the slide is adequate for the diagnostic interpretation.
 Participation in external programs also contributes valuable
information for a quality assurance program.
. There are two distinct systems that can be used to deliver
quality assurance such as selective system where stained
preparations from departmental archival records are used to
assess the quality of staining or distributive system in which
participating laboratories are asked to stain sections that have
been submitted by the scheme organizer.

Quality control and assurance plan for
histopathology lab

 Designing a quality control and assurance plan in histopathology

should focus on three elements: (1) preanalytical phase,
 (2) the analytical phase and
 (3) the post-analytical phase as defects may occur at any of
these phases, resulting in an erroneous diagnosis.
Pre-analytical aspects

 It includes sample collection, transport, accession and tissue

processing and submission of the slide for reporting.
 Studies have indicated that errors related to pre-analytical phase
can endanger quality of histopathology report.
 The accuracy of the final diagnosis is a measure of the
effectiveness of many elements including specimen collection,
gross dissection and section, tissue processing, embedding,
sectioning and staining
 Patient or specimen identification is one of the most important
aspects and it begins with specimen labeling and accessioning.
 Other critical element is adequate clinical history and has been
shown to affect the accuracy and completeness of pathology
reports. Standard operating procedure for sample accession,
identification, rejection of the sample and all steps of tissue
processing must be documented and displayed in laboratory and
the technical staff should be aware of its contents.
 Precise and systematic gross description, dissection and
selection of sections for microscopic study are crucial parts of the
pathologic examination and often cannot be remedied if omitted
or done poorly at the time of the initial work up.
 The stages of fixation, dehydration and clearing must be of
sufficient length to ensure completeness.
 All the equipment and instruments used in the laboratory should
be of standard quality and calibrated at periodic intervals.
 Disposable blades can provide a sharp cutting edge from which
flawless 2-4 μm sections can be cut with ease.
 The microscope and its parts should be serviced regularly to help
in imparting an accurate diagnosis.
 In case of frozen section, it is critical that the pathologist
communicates effectively with clinicians to verify clinical history,
appearance and exactly what information the surgeon wants.
 Even though overall diagnostic accuracy of frozen section ranges
from 89% to 98% in various studies, frozen artifact can produce
inferior slides for microscopic examination and sampling errors
can result from the heterogeneity of a tumor.
 Checklists to ensure quality control during pre-analytical phase
has been provided below.
Checklists to ensure quality control during
pre-analytical phase

 1. Sample collection and transport

 Biopsy
 - Appropriate procedure with adequate size and depth of the
tissue
 - Specimen from representative area, avoid mechanical trauma
 -Label specimen properly
 -Handle specimen gently
 Fixation
 - Immediate primary fixation
 - Suitable fixative in adequate quantity (ratio of at least 20:1) is
used in a container of an appropriate size.
 - Check fixative pH
 -Expedite large specimen fixation
 - Avoid unnecessary delays
 2. Referral form
 - Adequate clinical history
 - Pre-designed referral form should be made available to all
points of specimen collection
 3. Accession by laboratory
 - Entering details into a log book
 - Assigning a unique laboratory number
 4. Gross analysis
 - Check fixation status especially for large specimen
 - Precise and systematic gross description
 - Prepare thin slides
 - Adequate sampling
 - Careful sampling of specimens, avoid cross contamination
 - Choose appropriate cassettes and clearly label
 - Care is taken to avoid traumatizing delicate specimens,
particularly those that are incompletely fixed (handle carefully,
do not crush, always use sharp blades)
 - Cassettes are never overloaded with tissue thus allowing ready
access to processing reagents and preventing distortion of
specimens. If the volume of tissue is too great a second cassette
is used.
 5. Tissue processing
 - An appropriate schedule is chosen for the tissue type and size
 - Sufficient time to ensure completeness
 - Avoid prolonged contact with the reagents of tissue processing
 - Maintain reagent quality and use high quality wax
 - Planned changing of chemicals used for processing based on
the number of tissues passed through
 6. Embedding
 - Recording the temperature of paraffin wax bath, embedding
center hot plate, water floatation bath and slide warming table
daily.
 - Orientate the specimen carefully
 - Choose an appropriate mold
 - Handle specimen gently
 - Avoid excessive heat
 - Don’t overfill molds
 7. Cutting of sections
 - High quality, sharp blades are always used for cutting
 - Replacing microtome knives by disposable blade
 - Optimise knife tilt angle
 - Avoid freezing damage
 - Carefully trim blocks
 - Servicing and periodic calibration of the microtome
 - Use cold blocks - warm block may result in excessive
compression of the sections
 - Cut sections slowly
 8. Floatation
 - The water in the flotation bath is replaced regularly, use clean
water and check water temperature.
 - Avoid Cross-contamination
 - Ensure slides are clean
 -The water surface is always skimmed between specimens to
avoid contamination of one section with cells from another.
 - Don’t float from multiple blocks together in a floating bath
 - Avoid Wrinkles in Sections
 - Avoid over expending sections
 - Don’t damage floating sections, extreme care is taken to avoid
damaging floating sections when mechanically removing wrinkles
with a brush or forceps.
 - Prevent bubbles under sections, care is taken to avoid the
formation of air bubbles in the
flotation bath.
 Any visible bubbles are dislodged before the sections are laid on
the water.
 9. Section Drying
 -Sections are drained briefly before being placed in the slide drier
or onto a hotplate.
 - Dry for appropriate time, the minimum and maximum slide
drying time is monitored.
 10. Staining of slides, mounting and labelling
 - Each step in the staining protocol is accurately timed.
 - Ensure complete dewaxing
 - Standardized staining protocols for routine and special stains.
Agitation, wash and drain times are optimized for all steps during
staining.
 - Monitor Haematoxylin quality and eosin pH
 - Ensure complete nuclear blueing
 - Renew Reagents Regularly. Solvents and staining reagents are
regularly replaced based on the number of slides stained or racks
processed.
 - Thoroughly dehydrate before clearing and coverslipping.
 - Renew Reagents Regularly. Solvents and staining reagents are
regularly replaced based on the number of slides stained or racks
processed.
 - Thoroughly dehydrate before clearing and coverslipping

 Store Reagents Correctly. Some require refrigeration because

they are inclined to support the growth of fungi or molds. Others
are light sensitive and require storage in the dark.
 11. Immunohistochemistry stain
 - Always use appropriate positive and negative controls that are
carefully examined to validate results. Internal positive and
negative controls are also important and provide an excellent
means of ensuring quality assurance in IHC.
 - Avoid Section Adhesion Problems. Avoid the use of protein-
based section adhesives in the flotation bath (glue, starch, or
gelatin), particularly on charged slides.
 Avoid Concentration Gradients. Concentration gradients are
avoided by careful application of reagents.
 - Avoid Background Staining. Appropriate protein block is always
used.
 - Standardize Washing Steps. Use standardized washing steps
throughout (duration, volume and form of agitation). This will
ensure consistency of results.
 12. Insitu Hybridization
 - Use High Quality Sections. Take particular care to use thin, flat
sections that have been
thoroughly dried onto the slide. Use charged slides for ISH.
 - Choose Probe Carefully. Choose your probe carefully with regard
to its sensitivity and
specificity.
 - Optimize Pretreatment Conditions. Choose appropriate
pretreatment and optimization conditions. These will depend on
fixation and tissue type.
 - Use Appropriate Detection System. Choose a sensitive detection
and visualization system and optimize incubation conditions.
 - Standardize Washing Steps.
 Use standardized washing steps throughout (duration, volume
and form of agitation).
 This will ensure consistency of results.
 13. Equipment
 Equipment and instruments used in the laboratory should
be standard quality
 Equipment and instruments should be calibrated at
periodic intervals
 The microscopes and its parts should be serviced
regularly
 14. Submission for reporting
Analytic aspects

 The analytical phase is related to slide reading along with

relevant data and preparation of report. Pathologist must have
judgment, be conscious of the patient's welfare and should
always strive to provide accurate diagnosis.
 He/she must constantly seek out new knowledge and adopt new
practices as they become available.
 Assessment of analytical aspects in histopathology is not easy
because of the subjectivity of the reports. Intradepartmental
discussions, comparison with other reports (frozen, cytology, or
histopathology), blinded random case review, external
consultations and review by experts are some of the steps which
can be taken to improve the quality.
 Complete reporting in pathology has been given sufficient
attention in recent times particularly in oncology, where many
protocols are dependent on the pathological staging of tumors.
Post-analytical aspects

 Post-analytical phase involves the activities that follow the

analytical phase including preparation and delivery of the report,
archiving of request form and report, storage of the reported
specimen for set retention period and safe disposal of specimen
thereafter.
 All slides and paraffin blocks should be stored indefinitely if
facilities are available as these are of permanent nature and can
be evaluated on need.
 Laboratories should attempt to achieve the shortest turnaround
time. The retention period for specimens has always been a
subject of debate and national guidelines for this are warranted.