The best analytical techniques provide results that are only as meaningful as the quality of the specimen that has been submitted for analysis.
The quality of any laboratory test result is dependent on many variables, the first of which begins before the sample arrives at Pathology Lab.
Specimen handling includes best practices in collection, packaging and transport to the laboratory in a timely manner and under optimal environmental conditions that will not compromise the integrity of the specimen.
A quality diagnosis in pathology is directly related with three phases:
Pre-analytical phase consists of the patient preparation, collection, handling, packaging and shipping of the biological specimen before the arrival at the laboratory. That is, it encompasses all activities that precede laboratory techniques and procedures.
Analytical phase begins with the execution of the procedures and techniques implemented in the laboratory for sample processing, through internal quality control, and ends with the delivery of slides to the pathologist.
Post-analytical phase begins after the microscopic visualization of the slides and when the pathological report is completed, and delivered to the patient.
Proper Identification of Specimens
The participation of all professionals involved in the collection, packaging and sending of samples to the pathology lab is fundamental, so that the final diagnosis can be rigorous and provide the patient with adequate and accurate treatment.
Any error in user and / or sample identification can lead to serious misdiagnosis.
Specimens must be accompanied by a paper requisition, prepared either by hand or printed from an electronic ordering system. The requisition, at a minimum should contain the following information:
- Adequate patient identification information (e.g., name, address, telephone number, medical record number;
- Patient gender;
- Patient date of birth, or age;
- Name and address of physician ordering the test;
- Test(s) requested;
- Date of specimen collection, when appropriate;
- Source and type of specimen and time of collection, when appropriate;
- Clinical information, when appropriate
- Ensure that all specimen container caps and lids are properly tightened to prevent leakage
- Properly complete the requisition
- Collect the specimen(s) and transfer to a proper transport container, if needed
- Place on the container so that the label does not cover the handwritten patient name
- Frozen specimens must be transported in insulated containers surrounded by an ample amount of dry ice to keep the specimen frozen until it reaches the laboratory
- If the specimen has been classified as an “infectious substance,” transport in a box designed to meet the legal requirements.
Surgical Pathology Exams
- Frozen sections
1. Frozen sections
Biological (cytological or histological) specimen examined during a surgical procedure to determine the nature of a tissue or lesion or to establish the determination of the surgical margin.
There are two major types of specimens submitted for surgical pathology analysis: biopsies and surgical resections.
A biopsy is a small piece of tissue removed primarily for the purposes of surgical pathology analysis, most often in order to render a definitive diagnosis.
Biopsies correspond to small single or multiple fragments, usually irregularly shaped and without characteristic gross appearance. This category includes endoscopic gastrointestinal tract biopsies, transbronchial biopsies, bladder or prostate transurethral resection products, core needle biopsies of the breast, prostate, liver, kidney, skin punch, cervical biopsies, endometrium curettage products, among others.
Very small biopsies may be placed on filter paper prior to introduction into the vial. This procedure ensures a better orientation of the fragments, which facilitates their subsequent manipulation in the laboratory.
Surgical resection specimens
Surgical resection specimens are obtained by the surgical removal of an entire diseased area or organ. Some exemples include partial or total organ resection, large neoplasms and limb amputation products.
These procedures are often intended as definitive surgical treatment of a disease in which the diagnosis is already known or strongly suspected. Pathological analysis of these specimens is critically important in confirming the previous diagnosis, staging the extent of malignant disease, identifying the presence of unsuspected concurrent diseases, and providing information for postoperative treatment.
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Although the ideal fixative is far from being found, 10% buffered formaldehyde is the most commonly used fixative for most histological specimens in surgical pathology.
Specimens should be fixed in 10% buffered formalin immediately after collection to minimize the effects of autolysis. The ideal formaldehyde to tissue specimen ratio is of 10 volumes of formaldehyde to one specimen volume. When the specimen is too large to reach the proper ratio, it should be promptly shipped to the laboratory to minimize the effects of autolysis. As long as the specimen has a fixative, do not put the sample in the refrigerator.
3. Cytological Specimens
From an epidemiological point of view, cervical cytological examination is one of the most important examinations, as it contributes significantly to reduce the cervical cancer mortality rate. For good diagnostic acuity the specimen must be of good quality and this quality relys on three factors: collection, fixation and transport procedures to the laboratory.
Cervical cytology examinations may be performed conventionally or by methods known as liquid based cytology. The result is provided through interpretative reporting with the nomenclature recommended by the Bethesda System.
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Collection date, last menstruation date, hormonal status (eg pregnancy, postmenopause), contraceptive method used, previous history of intraepithelial neoplasia, cervical or extra-genital cancer, history of systemic chemotherapy, history of pelvic radiotherapy, history of gynecological surgery, cryosurgery or electrocauterization, abnormalities present at gynecological examination, risk factors for cervical cancer (eg sexually transmitted disease, early sexual activity, number of pregnancies).
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Pap smears (glass slide cytology) should be fixed immediately after collection to avoid drying artifacts that impair diagnosis.
Immediately after pap smear collection, fix the cells with fixative sprays (eg hairspray), trying to completely cover the area containing the smear. The smear and spray should be at a distance of 15 to 25 cm (if the smear is too close, the hairspray jet could spread the smear. On the other hand, if it is too far away the specimen will not be fixed correctly. No need to put in the fridge.
Liquid based cytology. After specimen collection the brush should be immersed and “shaken” in the bottle containing the cell preservation solution. Depending on the method used, the brush remains or not inside the tube. No need to put in the fridge.
In this method, cells are collected after they have been either spontaneously shed by the body (“spontaneous exfoliation”), or manually scraped/brushed off of a surface in the body (“mechanical exfoliation”).
Exfoliative cytology examination includes fluid and secretions (eg, urine, ascites, pleural, pericardial, lavage, and others).
Urine cytology specimens should be obtained as described in chapter urinary cytology.
To preserv exfoliative cytology specimens for short time period, put them in the refrigerator. Many samples remain relatively well preserved for up to 48 to 72 hours in a refrigerator (2-8 °C) without fixation, especially when dealing with pleural fluids. Other liquids, such as urine and cerebrospinal fluid, may deteriorate after 24 hours even if stored in a refrigerator.
In order to avoid deterioration, add equal amount of 50% alcohol to the liquid and refrigerate. However, although this technique opposes the preparation of air-dried slides, it is very effective in detecting lymphomas and leukemias.
Fine-needle aspiration cytology (FNA) is a fast, non-painful and invasive exam that is very useful in the diagnosis of palpable and non-palpable lesions. In the case of non-palpable lesions, the aid of imaging methods is essential.
This examination is performed with a fine needle (usually 23-gauge) attached to a syringe to collect cells from lesions or masses in various body organs, often with the application of negative pressure (suction) to increase yield. Cells are then placed on glass slides following microscopic observation. As a rule, these smears air dry.
Cibas, E. S., & Ducatman, B. S. (2009). Cytology: Diagnostic principles and clinical correlates. Philadelphia, PA: Saunders/Elsevier.