cells of human origin (blood cells). Most of these cells are not usually found in the stool; however, they may be found in the presence of parasites and/or other disease-producing organisms. These cells may occur in the stool without the presence of a specific etiologic agent (allergic conditions, ulcerative colitis, etc.). The buffy coat cells (white blood cells [WBCs]) can also be used as a QC check for various fixatives. The WBCs normally found in the buffy coat portion of whole blood include polymorphonuclear neutrophils, eosinophils, lymphocytes, and monocytes (macrophages). When blood is visible in the stool (indicating the presence of fresh blood), the WBC morphology is very similar to that seen on the stained thin blood film used for a differential. However, if the blood has been in the gastrointestinal tract for some time, the cellular morphology changes slightly.
QC for Schaudinn’s Fixative
1. Collect a fresh tube of blood (lavender top, EDTA anticoagulant), or get a tube from the hematology section (high WBC count, if possible).
2. Centrifuge the blood, and remove the buffy coat (layer containing WBCs).
3. Add the buffy coat to approximately 2 to 4 g of fresh, soft stool, and mix thoroughly but gently.
4. This stool-WBC mixture can be smeared onto several slides and immediately immersed in Schaudinn’s fixative.
5. After staining, if the WBCs appear well fixed and typical morphology is visible, one can assume that any intestinal protozoa placed in the same lot number of Schaudinn’s fixative would also be well fixed, provided that the fecal sample was fresh and fixed within recommended time limits.
QC for Fixative containing PVA
1. Mix approximately 2 g of fresh, soft stool with 10 ml of fixative containing PVA (in solution, ready for use). The ratio of stool to fixative should be approximately 1/3 to 1/5; do NOT add too much stool to the vial.
2. To this fixative-PVA-stool mixture, add several drops of buffy coat cells (concentrated/collected as above) and mix gently.
3. After a 30-min fixation, pour a small amount of the fixative-stool-PVA-cell mixture onto a paper towel to absorb the excess PVA.
4. The material can then be smeared onto several slides, allowed to dry thoroughly (60 min at room temperature or 30 min at 35°C), and stained. After staining, if the WBCs appear well fixed and typical morphology is visible, one can assume that any intestinal protozoa placed in the same lot number of PVA fixative would also be well fixed provided that the fecal sample was fresh and fixed within recommended time limits.
Note Entamoeba coli mature cysts are very difficult to fix properly and may be difficult to identify on the stained smear. For this reason, it is possible to have fixatives that are adequately QC checked but do not always yield good morphology for this particular organism. Use of a longer fixation time (60 min) sometimes produces better morphology after staining.
Note The same approach can be used for other stool preservatives (sodium acetate-acetic acid-formalin [SAF], zinc- and copper-based fixatives, other single-vial collection systems, and the Universal Fixative/TOTAL-FIX).
QC for Stains (Trichrome, Iron Hematoxylin, etc.)
1. Slides may be prepared from a known positive fecal specimen preserved in PVA or SAF. These slides can then be stained and checked for typical organism morphology. Positive PVA-preserved samples can be purchased commercially; many slides can be prepared from one sample. QC slides (already prepared and ready for staining) are also available commercially.
2. Buffy coat cells (after appropriate fixation) may also be used to check the staining properties of each new lot number of stain.
Note Do not prepare too many slides in advance. If dry fecal smears containing PVA are held too long before staining, the organism or buffy coat cell morphology may be poor.
Instruments and Equipment
General requirements include the following: (i) all instruments and equipment should be on a routine preventive maintenance schedule, and records should be maintained in writing; (ii) instrument manuals and maintenance/service records should be available for technical personnel to review; (iii) all thermometers should be checked and calibrated against certified standard thermometers, and the information should be recorded in writing; and (iv) checked and calibrated thermometers should be in every refrigerator, freezer, incubator, water bath, heat block, etc., and temperatures should be recorded on each day of use. As specific requirements, (i) centrifuges should be calibrated for correct speed and the calibration should be recorded on the instrument, and (ii) all microscopes should be calibrated by using a stage and ocular micrometer, the calibrations should be posted on the instrument, and recalibrations should be performed if any ocular or objective is switched on that particular microscope. It is recommended that microscopes be recalibrated once each year, even if the oculars and objectives have not been changed. This is particularly true if the microscopes receive heavy use.
Reference Materials
Reference materials should be available for comparison with unknown organisms, refresher training, and the training of additional personnel. Ideal reference materials include formalin-preserved specimens of helminth eggs, larvae, and protozoan cysts; stained fecal smears of protozoan oocysts, cysts, and trophozoites; and positive blood smears. Color slides and atlases are recommended, although the level of microscopic focus cannot be changed. Reference books and manuals from a number of publishers are available, and selected ones should be part of the parasitology library. It is also recommended that you maintain a list of consultants who can be called in case questions arise.
Patient outcome measures often refer to aspects of quality assurance that are monitored in addition to narrower parameters such as QC. Among the issues that are appropriate to monitor on an ongoing basis are the following:
1. Submission of the appropriate specimen within the correct time frame for collection
2. Condition of the specimen when received by the laboratory (too old, volume not sufficient, contains interfering substances, etc.)
3. Turnaround time for test results
4. Appropriateness of STAT requests
5. Improvements in compliance with established guidelines for the above, particularly after continuing-education efforts
6. Clinical relevance of and documented use of test results
All of these quality assurance parameters can be monitored by using The Joint Commission (formerly JCAHO) 10-step criteria for developing and implementing monitoring and evaluation activities (57; also http://www.jointcommission.org/accreditation/lab_standards_information.aspx [accessed 5/26/2015] and http://www.jointcommission.org/assets/1/18/2012_DSC_Cert_Guide.pdf [accessed 5/26/2015]). These 10 steps are listed below.
1. Assign responsibility. The designated individual identifies and assigns responsibilities to others within the department and ensures that these responsibilities are fulfilled.
2. Delineate scope of care. Delineating the scope of care involves identifying the diagnostic and therapeutic modalities used, times and locations where services are provided,
