Principles of specimen collection in pathology

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Principles of specimen collection in pathology

• Jumphol Mitchai MD
• Department of pathology KKU

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Pathology services

• Surgical pathology
• Cytopathology
• Exfoliative cytology
• Body fluids
• Fine needle aspiration (FNA, FNAC, FNAB)
• Cytogenetics
• Autopsy

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Surgical pathology

• Examined all tissues and foreign objects removed
  from living patients
• Specimens not to be submitted must be written as
  hospital policy
• Practice of pathologists
• Pathological diagnosis Gold standard in
  medicine

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Specimen collection in surgical pathology

• Incisional biopsy for diagnostic purpose
• Excisional biopsy for diagnostic and
  therapeutic purposes
• Tissue collection by instruments needle
  biopsy, core biopsy, punch biopsy, curettage,
  shave biopsy, Bard biopsy, endoscopic biopsy
• Necropsy biopsy from dead body
• Organs from operation rooms

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General rules for the biopsy

• Larger the lesion - more numerous biopsies
• Ulcerated tumor recommend peripheral biopsy
• Avoid necrosis and hemorrhage areas
• All fragments must be sent to the pathology
  department
• Not crushing, squeezing, cautery
• Immediately fix in fixatives
• Orientating specimen e.g. deep margin, superior
  and inferior margins by using sutured threads
  (long, short)

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Containers

• Transparency
• Lid
• Leak-proof
• Wide mouth, flat bottom
• Size corresponding to specimen volume
• Label

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Fixation of the tissue

• Hard tissues take more time to thoroughly fixed
  than soft tissue.
• Penetration rate, around 1 mm./hr. in general
  (variable, depends on consistency of the tissue).
• 10 times volume of fixative over the specimen
  volume is needed.

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Fixatives

• Neutral buffered formalin - fixation time 12-24
  hours.
• Formalin (40 aqueous solution of formaldehyde) -
  100ml
• Sodium dihydrogen orthophosphate (monohydrate) -
  4g
• Disodium hydrogen orthophosphate (anhydrous) -
  6.5g
• Distilled water - 900ml
• This fixative is suitable for most histological
  purposes. It is to be preferred to formol-saline
  (a single 10 solution of formalin in 9 aqueous
  NaCl) as formalin pigment is avoided. Specimens
  may be stored in this fluid. The solution is
  isotonic.

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• specimen(organ), method of specimen collection,
  characteristics, clinical informations)

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Filling the request forms (1)

• 1. identified of the patient
• 2. identified specimen ( gross appearance, cyst,
  vascular, inking for margin) type of
  surgery ( biopsy, wide excision) surgical
  finding
• 3. adequate clinical history (necessary)
• prior diagnosis, immune status (HIV),
  prior treatment radiation, chemotherapy bone
  lesion X-ray finding

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Filling the request forms (2)

• bone lesion X-ray finding
  hematolgic disease CBC, bone marrow
• female reproductive lesion LMP, hormone
• hepatitis LFT4. Clinical diagnosis or
  differential diagnosis

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Immunohistochemistry and HER2 study

• Use formalin-fixed paraffin embedded tissue
  (paraffin block)
• Must have request forms

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Frozen section

• Intraoperative consultation
• Fresh tissue was immediately freezed in liquid
  nitrogen and make an ultrathin section in
  cryostat
• Frozen section is not a permanent section

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Cryostat

• Cryostat microtome in the freezing cabinet.

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Frozen sectioning
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Indications for frozen section

• Benign or malignant lesions
• Adequacy of surgical margins
• Metastatic lymph node sentinel nodes in breast
  carcinoma
• Presence or absence of ganglion cells in large
  intestinal wall of Hirschsprung disease

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Diagnosis of frozen section

• Benign VS malignant not specific diagnosis
• In case of doubt wait for permanent section

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Specimens for frozen section

• Fresh tissue
• lt 1cm. in size is recommended.
• Put in plastic bag, tightly closed with rubber
  bands
• Label on the plastic bag
• Place the plastic bag in ice
• Send it to pathology lab immediately

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Inappropriate specimens for frozen section

• Large tumor or specimens
• Highly infectious specimens TB, HIV
• Follicular lesion of thyroid (can not diffentiate
  between benign and malignant)

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Cytopathology

• The study of individual cells or small clusters
  of cells for diagnosis and screening of diseases,
  including cancers
• practice of screeners, cytotechnicians,
  cytopathologists

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Specimen collection in cytopathology

• Specimens
• Smear slide
• Cervix (Pap smear)
• Fine needle aspiration (FNA)
• Bronchial brush
• Fluid
• Sputum
• Bronchial wash
• Body fluids, urine, CSF

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Cytogenetics

• Study structure of chromosome from cell culture
• FISH in chromosome of leukemia
• Specimens
• Whole blood
• Bone marrow
• Amniotic fluid

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Specimen collection for cytogenetics

• Sterile techniques
• 2-5 ml. of blood or bone marrow in heparinized
  tube or sterile heparinized syringe
• 20 ml. of amniotic fluid in sterile disposable
  syringe
• Send to lab immidiately (with request form) or
  keep in refrigerator not more than 12 hours

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Service hours for cytogenetics

• For blood specimen Mon, Tue, Fri 9.00-16.00
• For Bone marrow Mon, Thur 9.00-16.00
• For amniotic fluid Tue 9.00-15.00
• Blood and bone marrow for FISH Mon-Fri
  9.00-16.00

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Thank you