Table 2 Summary of actions and recommendations for the pre-analytical phase

Sample collection and conditioning

1.Personnel responsible for specimen collection and for completing the request form with clinical information: physician, surgeon, or radiologist

2.Personnel responsible for registering information regarding the time of specimen collection and the time of cold ischemia (defined as the time between tissue removal from patient until placement into the fixative): nursing team (operating room) or the radiology assistant/ technician (radiology services)

3.The excised material must be clearly specified in the request form and checked by the nursing team before placement in the containers with fixative

4.Handling of specimens before fixation:

• Outpatient procedures: keep in saline solution if fixation will not be performed immediately (for example, in cases that require radiography or photographic documentation of the specimen)

• Surgical specimens:

○ Small samples (nodulectomies, lymph nodes, lumpectomy), measuring less than 5.0 cm or at physician discretion, can be immediately placed in the fixative, fully submerged

○ Larger samples, such as mastectomies and wide local excisions, should be sliced in case they are not immediately sent to the pathology laboratory (see below for details)

○ Samples that had undergone an intraoperative frozen section should be sent fresh to the pathologist, who will be responsible for the specimen manipulation until the intraoperative diagnosis. After the test, the specimen will follow the same workflow described for samples that are not submitted to intraoperative procedures

5.Preparation of larger specimens

• Specimens with larger volume need to be properly prepared for adequate fixation. Although formalin is a good fixative, its action is slow, as it penetrates the tissue with a speed of 1 mm/hour at room temperature. This information can be used to support the choice of the thickness of the fragments (thinner thickness, in case delays in the specimen dispatch to the laboratory, for example, during the weekend or holidays). It is recommended that surgical specimens be cut in parallel slices performed from the deep fascia towards the skin, without transfixing the surgical piece so it can be recomposed in the laboratory. This procedure needs to be agreed between the pathology laboratory and the surgical team

• The pathologist is responsible for training the personnel involved in the procedure after the specimen excision, such as the surgical team members, technicians, paramedics etc., depending on the local conditions

• Ideally, before slicing, the resection margins should be identified and inked. In this case, it is necessary to dry the specimen using paper towel, apply the ink followed by acetic acid or vinegar so the ink can fix properly without dissolving in formalin and during the processing, thus allowing the proper assessment of the surgical margins

• Inadequate fixation impairs the histopathological diagnosis (differential diagnosis between benign and malignant, histological tumor typing and grading, and the immunoreactivity of target molecules

6.Specimen labeling and identification (nursing team)

• Labels for container or slide identification should be printed using computers or written in pencil in adhesive tape, and contain patient’s name and information about the specimen

• Ideal scenario: Bar-code or QR code

• The label should be placed on the primary container, not in the lid

• Certify that the received specimen matches the description provided in the medical request

7.Placement in the containers

• Containers should preferably be rigid, impermeable, break-resistant, and non-reactive to fixatives

• Previously identified by the nursing team

8.Fixation

• Register the time the specimen was placed in the fixative

• Recommended cold ischemia time: less than 1 h

• Recommended type of fixative: 10% neutral phosphate buffered formalin (40% formaldehyde diluted to 10%—elevation of pH to ~ 7)

• Fixative volume: 10 to 20 times the size of the specimen

• Fixation time of tumor samples recommended for hormone receptors and HER2: 6–72 h

Pathological exam request

• Responsibility of the medical team

• The request form must accompany the specimen during the complete journey, from collection to the end of pathological exam

• Should specify:

• Laboratory of destination

• Patient identification

• Clinical diagnosis/diagnostic hypothesis

• Summary of the clinical history

• Procedure performed

• Date of procedure

• The specimens should be preferably numerate and properly described regarding its type, laterality, and topography

• Type of test to be performed (e.g., immunohistochemistry, molecular tests)

Transportation to the pathology laboratory

• Forms of sending the specimen/material

• Intra-hospital transfer (the pathology laboratory is located in the hospital or clinic itself)

• Laboratory outside the hospital (transportation using messenger service or mail):

○ Adequate conditioning: primary container (container with the specimen properly identified), secondary (leak-proof) and tertiary (rigid, accompanied by the identification of the sender and the recipient, identification of the biological material, and phone number contact in case of accident)