Histology Exam Flashcards
What is cryotomy / frozen sectioning
A technique that uses a cryotome to prepare thin and frozen sections for biological tissues
What are frozen sections used for?
Can be used for tissue analysis that allows for rapid interpretation and diagnosis of tissue during surgery
Can also be used to prepare sections containing lipids and enzymes that can be easily lost in alcohol or paraffin sections
Describe the impact of common errors experienced in cyrotomy and what are the solutions to these errors
Inadequate tissue preparation can lead to poor sections and distorted morphology = sections must be well frozen and have adequate medium (OCT) before sectioning
Inappropriate cryostat temperature - incorrect temperature can cause variations in thickness and sectioning - temperature must be checked and regulated before sectioning
Tissue orientation - tissue must be as flat as possible on chuck to avoid diagnostic elements of the tissue being lost and allow for sectioning of full representation of structures
Clinical significance of cryotomy
Margin assessment - in cancer surgery (MOHs) cryotomy aids in assessing the margins of excised tumors. It helps surgeons ensure that cancerous tissue is completely removed while preserving healthy tissue.
Diagnostic pathology - used to obtain urgent frozen sections during surgery. provides rapid on-site evaluation of tissue samples to guide surgical decisions.
Tissue processing steps in order
Fixation
Dehydration
Clearing
Impregnation
Embedding
Describe the process of Dehydration
Since melted paraffin wax is hydrophobic most of the water in a specimen must be removed before it is infiltrated with wax. This is done by immersing specimen in a series of increasing alcohol concentrations until pure water free alcohol is reached.
Describe the process of Clearing
Although the tissue is water free now, wax cannot infiltrate as wax and alcohol are not immiscible hence a clearing agent is used to displace the alcohol in the tissue. A common clearing agent used is xylol
Describe the process of Impregnation
The tissue can now be infiltrated by a suitable histological wax eg. Paraffin, resin. Wax infiltration provides tissue with structural support and integrity and allows sections as thin as 2uM to be cut and retain elasticity to flatten fully on a warm water bath.
Role of tissue processing
Processing involves dehydration, clearing and infiltrating the fixed tissue in a support medium (usually paraffin) to make it suitable for sectioning and staining.
Role of fixation and clinical significance
Fixation involves preserving the tissue structure and cellular components while preventing processes such as autolysis and putrefaction. It coagulated proteins, stabilizes membranes and cross links cellular components.
Clinical significance
- Preservation of tissue morphology - adequate fixation and processing preserves the overall morphology of tissues
- Consistent tissue quality and preventing artefacts - consistent tissue quality is crucial for proper and accurate diagnosis. Inadequate fixation can lead to artefacts being produced which can lead to misinterpretations and misdiagnosis.
Role of embedding and tissue orientation
Embedding is the processing of surround tissue specimen with a suitable medium (paraffin wax) to provide support and facilitate thin sectioning for microscopic examination.
Tissue orientation refers to proper alignment and positioning of tissue specimen during embedding.
Common errors of embedding and solutions
Tissue folding - properly flatten and align tissues during embedding
Air bubbles - use gentle tapping or re-embed to remove / avoid bubbles
inadequate support - ensure mold is filled with enough wax
multilayers of wax in mold - re-embed and embed tissue quicker before initial wax inserted hardens.
Clinical significance of correct tissue orientation and important of adherence to blocking instructions
Accurate diagnosis - correct orientation ensures sections contain structures of interest for diagnosis
Margin assessment - correct orientation allows for accurate evaluation of margins during cancer / lesion specimens
Consistency and reproducibility - adhering to blocking instructions ensures consistent and reproducible results across different tissue samples
Avoidance of artifacts - avoiding artefacts such as poor morphology; inconsistent sectioning
Implication of multiple pieces of tissue on different planes in one cassette
Regions of diagnostic importance being missed and not present during pathological review.
Implications of Incorrect orientation of a skin punch biopsy
Implications include; distorted architecture, incomplete assessment, incorrect depth assessment and artifacts and misinterpretations.
The orientation of the specimen is crucial in dermatopathology as it helps determine the relationship between the epidermis, dermis and subcutaneous tissue as well as the location of specific structures or lesions within the skin layers
Microtomy principle
Sample preparation, Sectioning instrument, Sectioning technique, Section collection and Section handling and preservation
The principles of microtomy aim to achieve thin, uniform sections that preserve tissue morphology allowing accurate analysis, and facilitating subsequent staining and analysis. Adherence to these principles is crucial for high-quality sections for pathological examination and diagnosis.
Importance of good microtomy technique
Good microtomy technique is essential for obtaining high-quality sections, accurate diagnosis, efficient workflow and occupational health and safety. Adhering to proper microtomy techniques and following established guidelines and protocols ensures the integrity and quality of histological sections, enabling accurate analysis and interpretation of tissues for various applications.
Microtomy artefacts and remedies (8)
- Ribbons / curved sections = trim block until parallel, replace / move blade, trim excess paraffin
- Chatter = clean microtome/ tighten clamps, reduce angle for knife, use softening agent
- Sections not ribboning = adjust knife angle, replace blade, dull blade softly with brush
- Excessive compression - cool block / replace blade
- Sections expand on waterbath = reduce waterbath temperature
- Hard tissue / won’t cut - decrease cutting speed/thickness
- Sections rolling on the knife - replace blade / reduce section thickness
- holes/scores in section - knick in knife edge - replace the blade
Temperature of waterbath
40-50 C
QC definition and aim
QC is a measure of how well the measurement system reproduces the same result over time and under varying operating conditions. It is a system for verifying and maintaining a desired level of quality in an individual test or process
QA definition and aim
QA is a coordinated system designed to detect, control and prevent the occurrence of errors. It is a systematic monitoring of quality control results and quality practice parameters to ensure that all systems are functioning in a manner appropriate to excellence in health care delivery
it aims to further a clinician’s ability to appropriately care for their patients.
Lab Qc is designed to
Lab QC is designed to detect reduce and correct deficiencies in a laboratory’s internal analytical process before releasing patient results in order to improve the quality of results reported by the laboratory
Benefits of specimen tracking system
- Enables audit of samples from start to finish
- Creates a robust system that tracks and verifies the identity of every specimen at every point of the workflow
- Nothing is left to chance, everything is checked and confirmed; all steps are covered
- It eliminates human identification errors
- includes checks to ensure any potential problem is identified before there is a risk to patient safety.
Benefits of QC
Patients assured of receiving quality service from lab
Consistent results produced all the time
Internal processors are constantly checked
Types of QC
Internal and External
Types of Audits
Internal
External
Horizontal
Vertical
Examination
Horizontal audit
Examines one element in a process on more than one item
eg. specimen accessioning
Vertical audit
Examines multiple elements in a process, on one item
eg. following the path of the specimen from receipt to reporting
Examination audit
Examines a person undertaking a test procedure
Ensure that the person’s actions reflect SOP
Ensure the person understands the procedure
