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Principles of specimen
collection in pathology
Jumphol Mitchai MD
Department of pathology KKU
Pathology services
• Surgical pathology
• Cytopathology
– Exfoliative cytology
– Body fluids
– Fine needle aspiration (FNA, FNAC, FNAB)
• Cytogenetics
• Autopsy
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 ”
Specimen collection in surgical
• 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
General rules for the biopsy
• Larger the lesion - more numerous biopsies
• Ulcerated tumor – recommend peripheral
• Avoid necrosis and hemorrhage areas
• All fragments must be sent to the pathology
• Not crushing, squeezing, cautery
• Immediately fix in fixatives
• Orientating specimen e.g. deep margin, superior
and inferior margins by using sutured threads
(long, short)
Wide mouth, flat bottom
Size corresponding to specimen volume
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.
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.
การเตรียมสิ่ งส่ งตรวจ
1 การติดสลากบนภาชนะใส่ ขอ้ มูลผูป้ ่ วยให้ครบถ้วน ชื่อสกุล HN เพศ
อายุ แพทย์เจ้าของไข้ หอผูป้ ่ วย
2 การเขียนใบนาส่ ง
ใส่ ขอ้ มูลผูป้ ่ วยในทุกช่องรายการข้อมูล
specimen(organ), method of specimen
collection, characteristics, clinical informations)
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
Filling the request forms (2)
bone lesion : X-ray finding
hematolgic disease : CBC, bone marrow
female reproductive lesion : LMP, hormone
hepatitis : LFT
4. Clinical diagnosis or differential diagnosis
Immunohistochemistry and HER2 study
• Use formalin-fixed paraffin embedded tissue
(paraffin block)
• Must have request forms
Frozen section
• Intraoperative consultation
• Fresh tissue was immediately freezed in liquid
nitrogen and make an ultrathin section in
• Frozen section is not a permanent section
• Cryostat = microtome in
the freezing cabinet.
Frozen sectioning
Indications for frozen section
• Benign or malignant lesions
• Adequacy of surgical margins
• Metastatic lymph node : sentinel nodes in breast
• Presence or absence of ganglion cells in large
intestinal wall of Hirschsprung disease
Diagnosis of frozen section
• Benign VS malignant : not specific diagnosis
• In case of doubt – wait for permanent section
Specimens for frozen section
• Fresh tissue
– < 1cm. in size is recommended.
• Put in plastic bag, tightly closed with rubber
• Label on the plastic bag
• Place the plastic bag in ice
• Send it to pathology lab immediately
Inappropriate specimens for frozen
• Large tumor or specimens
• Highly infectious specimens : TB, HIV
• Follicular lesion of thyroid (can not diffentiate
between benign and malignant)
• The study of individual cells or small clusters of
cells for diagnosis and screening of diseases,
including cancers
• practice of screeners, cytotechnicians,
Specimen collection in cytopathology
• Specimens
– Smear slide
• Cervix (Pap smear)
• Fine needle aspiration (FNA)
• Bronchial brush
– Fluid
• Sputum
• Bronchial wash
• Body fluids, urine, CSF
วิธีการเตรี ยมสิ่ งส่ งตรวจทาง cytopathology
1 สิ่ งส่ งตรวจที่ป้ายบนสไลด์ให้แช่ใน95%Alc. ทันที
2 Fluid specimen ให้นาส่ งห้องlabทันที หรื อเก็บในตูเ้ ย็นได้
3 เขียนใบนาส่ งที่มีรายละเอียดของผูป้ ่ วย และประวัติการเจ็บป่ วย
4 ติดสลากระบุขอ้ มูลผูป้ ่ วยบนภาชนะหรื อสไลด์
5 ส่ งห้องปฏิบตั ิการทันที
• Study structure of chromosome from cell
• FISH in chromosome of leukemia
• Specimens
– Whole blood
– Bone marrow
– Amniotic fluid
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
• Send to lab immidiately (with request form) or
keep in refrigerator not more than 12 hours
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
เกณฑ์ การปฏิเสธสิ่ งส่ งตรวจ
ชื่อสกุล และHN ไม่ชดั เจน
ชื่อสกุลในใบนาส่ งและบนภาชนะไม่ตรงกัน
ไม่ระบุหน่วยงานนาส่ ง, แพทย์เจ้าของไข้
ไม่มีประวัติผปู ้ ่ วยและผลการวินิจฉัยทางคลินิก
ชนิดของสิ่ งส่ งตรวจไม่ตรงกับใบนาส่ ง
สภาพสิ่ งตรวจไม่เหมาะสม
Thank you