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TJUHBB

Tissue Harvesting

Standard Operating Procedure (SOP 06.002)

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PURPOSE

Tissue samples are collected from patients that have been through the informed consent process and agreed to participate in the tumor biorepository program. Tumor tissues are obtained by qualified personnel and are collected for the biorepository only if present in excess to that required for pathological assessment and diagnosis. The purpose of this document is to outline standardized procedures for the TJUH Brain Biorepository to follow during tumor tissue harvesting.

SCOPE

This standard operating procedure (SOP) describes how tissues should be harvested. The SOP does not cover detailed safety procedures for handling Human Biological Materials (HBMs) and it is recommended that personnel follow institutional biosafety guidelines.

REFERENCE TO OTHER TJUHBB SOPS OR POLICIES

  • TJUHBB POL 03.004 Personnel Training and Records
  • TJUHBB POL 02.002 Laboratory Safety Policy
  • TJUHBB POL 02.001 Chemical Hygiene Plan
  • TJUHBB POL 01.001 Quality Management Plan
  • TJUBB SOP 06.003 Snap Freezing
  • TJUHBB SOP 06.001 Tissue Collection and Transportation

ROLES AND RESPONSIBILITIES

This SOP applies to all qualified tumor biobank personnel and clinical staff at the collection centers that are involved in recruiting patients and the acquisition of informed and voluntary consent. This may include the following personnel:

Tumor Biobank PersonnelResponsibility
Attending Clinician (either neurosurgeon and/or neuropathologist)Identification and diagnosis of tissue malignancy, grossing of tissue and/or resection of excess tumor tissue for the biorepository
Tumor Bank ManagerCollection, harvesting, processing and storage of biospecimens

MATERIALS, EQUIPMENT AND FORMS

The materials, equipment and forms listed in the following list are recommendations only and may be substituted by alternative/equivalent products more suitable for the site-specific task or procedure.

Materials & EquipmentQuantity
Insulated Ice Box1
Sterile Specimen ContainerDepends on Quantity
Sterile Disposable Forceps1
Sterile Disposable Scalpel2
Sterile Disposable Petri Dish1
Pre-filled, Pre-labelled 2.0 mL CryotubesDepends on Quantity
Small Liquid Nitrogen Container1

DEFINITIONS

See the TJUHBB Glossary.

PROCEDURES

This procedure is intended to ensure that tissue samples will be collected from consented participants in a safe and efficient manner while eliminating the risks of contamination. To facilitate the use of innovative genomic and proteomic techniques, banked tissue that has been adequately processed is vital to obtaining products with high integrity and quality.

Tissue Harvesting

  • Treat all tissue as potentially infectious.
  • Have all materials necessary for tissue processing prepared in advance in the BSL-2 hood.

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Warning

Ensure that the resected tissue never desiccates or is contaminated by surrounding tissue or other samples. If appropriate, change scalpel blades between dissecting tumor tissue and surrounding uninvolved tissue.

  • Place the tissue in a sterile petri dish using clean technique. Be sure not to touch any surfaces that will be touched by the tissue (i.e. the inside of the petri dish).
  • Slice the tissue with a clean scalpel. Always use a clean scalpel between tissue samples or between normal and tumor tissue.
  • Identify areas of frank cauterization and/or necrosis and exclude these areas from the tissue specimen by dissection.
  • Specimens should be dissected into aliquots of equal size, with a minimum size of approximately two cubic millimeters (roughly, half the size of a grain of rice).
  • Specimens should be no larger than seven (7) cubic millimeters to ensure rapid freezing.
  • If nine (9) aliquots are available:
    • One (1) aliquot should be formalin fixed and placed in the cryovial containing 2 mL of 10% neutral buffered formalin. This cryovial is designated with a clear or white cap.
    • Two (2) aliquots should be preserved with RNAlater and placed in the cryovial containing 1 mL of RNA later. These cryovials are designated with a red cap.
    • Six (6) aliquots should be snap frozen and placed in the empty cryovials. These cryovials are designated by a blue cap.
  • If the tumor amount exceeds nine aliquots at the largest aliquot size (6-7 cubic millimeters), then additional snap frozen specimens can be collected.
  • The blue cryovials designated for snap freezing preservation should be placed into baskets in the dewar.
  • The blue cryovials designated for snap freezing preservation should be placed into a cryogenic dewer and immediately placed into the liquid nitrogen container on hand (see Figure 1).
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Figure 1. Cryosticks in cryogenic dewer

  • Allow 5 minutes for freezing to be completed and remove the snap frozen aliquots and place them into the designated cryogenic storage boxes for permanent storage.
  • The cryovials designated for RNAlater preservation should be held at room temperature for four (4) hours and then snap frozen in the same manner and finally placed into the designated cryogenic storage box for permanent storage.

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Timing of Specimen Processing

Timing is critical. Ideally, no more than 30 minutes must elapse between the time of biopsy/resection and time of freezing of a given sample. Records must clearly document actual time period (in hours or minutes).

APPLICABLE REFERENCES, REGULATIONS AND GUIDELINES

REVISIONS

SOP IDDate RevisedAuthorRevision Summary
SOP 01.00107/01/2015Molligan, JeremyCreated

VERSIONS & APPROVAL

VersionCategoryApprovalDate
01.001AdministrationStephen Peiper, MD07/01/2015

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