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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.
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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 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
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)
Containers • Transparency • Lid • Leak-proof • Wide mouth, flat bottom • Size corresponding to specimen volume • Label
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.
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.
การเตรียมสิ่งส่งตรวจ 1 การติดสลากบนภาชนะใส่ข้อมูลผู้ป่วยให้ครบถ้วน ชื่อสกุล HN เพศ อายุ แพทย์เจ้าของไข้ หอผู้ป่วย มีการติดสลากบนทุกภาชนะ 2 การเขียนใบนำส่ง ใส่ข้อมูลผู้ป่วยในทุกช่องรายการข้อมูล 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 : LFT4. Clinicaldiagnosis 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 cryostat • Frozen section is not a permanent section
Cryostat • Cryostat = microtome in the freezing cabinet.
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
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 bands • Label on the plastic bag • Place the plastic bag in ice • Send it to pathology lab immediately
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)
Cytopathology • The study of individual cells or small clusters of cells for diagnosis and screening of diseases, including cancers • practice of screeners, cytotechnicians, cytopathologists
Specimen collection in cytopathology • Specimens • Smear slide • Cervix (Pap smear) • Fine needle aspiration (FNA) • Bronchial brush • Fluid • Sputum • Bronchial wash • Body fluids, urine, CSF
วิธีการเตรียมสิ่งส่งตรวจทาง cytopathology 1 สิ่งส่งตรวจที่ป้ายบนสไลด์ให้แช่ใน95%Alc. ทันที 2 Fluid specimen ให้นำส่งห้องlabทันที หรือเก็บในตู้เย็นได้นาน12-24ชม. 3 เขียนใบนำส่งที่มีรายละเอียดของผู้ป่วย และประวัติการเจ็บป่วย 4 ติดสลากระบุข้อมูลผู้ป่วยบนภาชนะหรือสไลด์ 5 ส่งห้องปฏิบัติการทันที
Cytogenetics • Study structure of chromosome from cell culture • 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 syringe • 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 9.00-16.00
เกณฑ์การปฏิเสธสิ่งส่งตรวจเกณฑ์การปฏิเสธสิ่งส่งตรวจ ชื่อสกุล และHN ไม่ชัดเจน ชื่อสกุลในใบนำส่งและบนภาชนะไม่ตรงกัน HNไม่ตรงกับข้อมูลในเวชระเบียน ไม่ระบุหน่วยงานนำส่ง, แพทย์เจ้าของไข้ ไม่มีประวัติผู้ป่วยและผลการวินิจฉัยทางคลินิก ชนิดของสิ่งส่งตรวจไม่ตรงกับใบนำส่ง สภาพสิ่งตรวจไม่เหมาะสม