histopathology in practice - sample reception, dissection, processing, embedding
DESCRIPTION
A talk about the daily operations of a histopathology laboratory - from specimen reception through to tissue embedding.TRANSCRIPT
Histopathology in Practice
Sample Reception, Dissection,Processing, Embedding.
Della ThomasSenior Biomedical Scientist
Independent Histopathology Services
Histology
The science concerned with the microscopic structure of tissues and organs in relation to their function.
Also called microanatomy.
Specimen Reception
Specimen Unpacking
Mickey MouseRt nasal polyp
Dr. Makewell26/11/1318/11/28DOB:
Specimen QC
MOUSEMr. MICKEY18/11/281 2 3 4 5 6Dr. Makewell26/11/13 8am
C/O loss of function - ability to sniffout cheeseO/E: nasal polyp? nature.
Rt nasal polyp
M
Specimen Labelling
Sample documentingAssign the
specimen to a pathologist
MOUSEMr. MICKEY18/11/281 2 3 4 5 6Dr. Makewell26/11/13 8am
C/O loss of function - ability to sniffout cheeseO/E: nasal polyp? nature.
Rt nasal polyp
M
Cassetting
Specimen Dissection - the stage
Dissecting props• Cutting board• Forceps• Ruler• Scalpel / knives / saw• Inks / dyes• Cassettes / lids• Biopsy pads / tissue• Filter bags• Weighing scales
Cut-up dictation
• “Morphology” computer system
• “Dragon” dictation system
• Hands-free
• Just scan the barcode and away we go!!
Specimen Categories
A Specimens only requiring transfer from container to tissue cassette.
B Specimens requiring transfer but with standard sampling, counting, weighing or slicing.
C Simple dissection required with sampling needing a low level of diagnostic assessment and/or preparation.
D Dissection and sampling required needing a moderate level of assessment.
E Specimens requiring complex dissection and sampling methods
Category A SpecimensBreast core biopsies
Endometrium
Colonic series
Category B Specimens
Cervical LLETZ
Small skin biopsy
Small lipoma
Category C Specimens
Appendix
Gallbladder
Haemorrhoids
Prepuce
Category D Specimens
Salivary gland tumour
Large Intestine (Crohn’s)
Skin with markers
Pigmented skin lesions
C/D
C/D
Category E Specimens
Testis (seminoma)
Thyroid (medullary Ca)
Breast cancer
Uterus (endomet. Ca)
Specimen Dissection
What does the pathologist need to know?
Provide good descriptions - say what you see!!• Shape• Colour• Texture• Dimensions• Weight• Distances from margin(s)• Orientation markers• Cut surface appearance etc....
• Keep your fingers crossed for good clinical history
Doctors’ handwriting!!
“dysplasia”?OR “lymphoma”?
Breast Ca....”Steroids”?OR “Stomach”?
“Nothing” suspiciousOR “Notably” suspicious?
Simple transfer
Prostate cores
Cervical biopsy
Specimen Sampling
• Literally, taking a sample of the tissue
• Representative
• Generally, fewer blocks required if the tissue looks uniform throughout (for benign cases)
• Sampling “rules”
Sampling rules: prostate chips• If the chippings weigh 12g
or less, the entire specimen must be processed
• If the chippings weigh more than 12g, a minimum of 6 cassettes must be processed
• For every 5g over 12g, one more cassette must be processed
Prostate chips (19g)
Sampled in 8 cassettes:
First 12g = 6 cassettes
19g = 7g over 12g
1 cassette per 5g over = 2 more cassettes.
Simple dissection
Specimen: Skin from back
Clinical details: Sebaceous cyst
Simple dissection
Unlock the pathologyFistula-in-ano
Unlock the pathology
Gallbladder
Unlock the pathologyPosterior view
Cystoprostatectomy
Inking• Resection margins
• Embedding instructions
• Orientation
• Distinguish between samples
• Identify the cut surface
• Acetic Acid
Inking resection marginsPigmented lesion
Cervical cone
Embedding Instructions
Ink dots instruct the embedder to embed the tissue a certain way.
Orientation - which way up?• Anatomical
Orientation - which way up?• Clockface
3 o’clock markerClinical Data:Suture marker at 3 o’clock.
“12 o’clock” markerClinical Data:Tag suture at 12 o’clock.
?
“12 o’clock” marker
Mapping on a larger scale
Red = RightGreen = Left
ProstatePosterior bladder
Anterior bladder
Ureter
Cystoprostatectomy
Cassette sizes
Calcified / firm tissue
• Femoral head
• Bone Marrow Trephine
• Ethmoid mucosa / nasal polyps (cartilage)
• Nail
• Hardened cysts
Most common:
Softening
• For bony / hard tissue: 10% Formic Acid
• For nail: Phenol or hair removal
cream.
Firm tissue testing methods• X-ray - Expensive / ? bench space, but
very accurate
• Chemical end-point test (Ammonium Hydroxide/Ammonium Oxalate) - very time consuming, but accurate
• Physical manipulation - not very accurate, may damage the specimen, but simple and inexpensive
Cutting instructions
• Levels• Special stains• Unstained Sections• Serial sections• Alopecia protocol• MM sentinel lymph node• Hirschprung’s protocol
Specimen Storage• Ventilated storage units
• Largest buckets lower shelves
• Units are in date (week) order
• 5-weeks’ worth of storage
• Only authorised specimens are discarded after 5 weeks
• Any outstanding cases are stored separately until further notice
ProcessingFormalin Alcohol Xylene Wax
Formalin
Alcohol
Xylene
Wax
Processing programs
EmbeddingThe immortalisation of tissue presentation
Embedding CentreMolten wax
Cold platesWax dispenser
Wax flow adjuster
Heated chambersUsed lids
Hot surface
Embedding tools
Embedding processDispense wax Align tissue
Cool in place Cassette on
Embedding processID bead Top-up wax
Cool plate
Leave to set
Embedding tips
• Ink dots: usually used to instruct the embedder to embed the tissue a specific way. Make sure you know your own lab’s protocol.
• Always keep your eye on the tissue: even for the most careful embedders, tissue can ping like tiddlywinks. Make sure you see where it lands!!
• Hide-and-seek: open lids / sponges carefully - tissue often sticks to them.~ why it’s important for the embedder to know number of bits in the cassette
• Cleanliness: always watch out for potential carry-over!!
Final block preparation
Imagination
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@dellybean (Della Thomas)
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Thank you