Q & A

May 2004

Richard A. Savage, MD, Editor

Q. Are there recommended procedures or policies for disinfecting the cryostat following frozen sections on a potentially transmissible infectious disease such as active tuberculosis, Coccidiodes immitis, or a virus that was not suspected clinically or on gross examination of the specimen? What is the appropriate timing for decontamination, and when is it appropriate to use a backup cryostat?

A. A cryostat is considered a high-risk instrument because it uses frozen, unfixed tissue that can contain viable infectious agents. Freezing the tissue does not inactivate infectious agents. In fact, question ANP.24250, which was added to the safety section of the anatomic pathology checklist to address these concerns, asks:

Is there a documented procedure for the routine decontamination of the cryostat at defined intervals and are decontamination records evident?

The explanatory commentary accompanying this question states that the interior of a cryostat should be decontaminated regularly with 70 percent ethanol. Trimmings and sections of tissue that accumulate inside the cryostat should be removed during decontamination. The cryostat should be defrosted and decontaminated with a tuberculocidal disinfectant at a time interval appropriate for the institution-once a week for instruments used daily. Freezing propellants under pressure is not advised because they may cause the splattering of droplets of infectious material.

The cryostat must be clearly marked as contaminated if a frozen section is performed on tissue from a patient known or suspected to be positive for HIV, hepatitis B or C, SARS-related coronavirus, prion disease such as Creutzfeldt- Jakob disease, or mycobacterial or systemic fungal disease. The cryostat must be decontaminated before further use. Laboratories, therefore, may need to have a backup instrument available.

Seventy percent alcohol has been shown to be effective against HIV and probably other viruses. To disinfect for tuberculosis or fungal disease, a lab should use a tuberculocidal or fungicidal disinfectant.

Bibliography

NCCLS. Protection of Laboratory Workers from Occupationally Acquired Infection; Approved Guideline—2nd Edition. NCCLS document M29-A2. Wayne, Pa.: NCCLS; 2001.

Thomas A. Merrick, MD
Department of Pathology
Presbyterian St. Luke’s Medical Center
Denver
Chair, CAP Safety Committee

Q. Our new owners are tardy in paying our suppliers, so we no longer have sedimentation rate controls. Is it acceptable to use a patient control from the previous day’s run? What range would be acceptable?

A. It is acceptable for a laboratory to retain a patient's sample and use it as quality control material for a clinical assay.

According to NCCLS, a patient specimen for the erythrocyte sedimentation rate assay may be used for quality control if the specimen is collected in EDTA, has a packed cell volume of 0.35 or less, and has an elevated ESR in the range of 15 to 105 mm/h. The specimen must be inverted well (16 times) before testing.

Acceptable specimen retention times are still in effect. For most laboratories, the ESR must be performed within four hours if kept at room temperature or 24 hours if refrigerated, as indicated by NCCLS. If these requirements are met, it is acceptable to use retained patient samples for QC material for this assay.¹

Reference

1. NCCLS. Reference and Selected Procedure for the Erythrocyte Sedimentation Rate (ESR) Test; Approved Standard–4th Edition. NCCLS document H2-A4. Wayne, Pa.: NCCLS; 2000. Vol. 20, No. 27.

Amy S. Gewirtz, MD
Ohio State University
Medical Center
Columbus
Member, CAP Hematology/
Clinical Microscopy Committee

Q. We occasionally receive cerebrospinal fluid from patients with ventriculoperitoneal shunts. The fluids from these patients do not look the same as other CSF samples. Should they be reported with the same normal ranges as CSF? If they should be reported with other ranges, can you provide several good references?

A. The cerebrospinal fluid in the ventricles is different from the cerebrospinal fluid in the lumbar region.¹ Small studies of ventricular taps versus lumbar taps in adults have demonstrated that protein levels in the ventricles are lower (mean, 15 mg/dL in ventricles versus 45 mg/dL in the lumbar area) and glucose levels and pH are five to 10 percent higher in the ventricles. Clumps of choroidal cells are observed more commonly in ventricular fluid than in lumbar fluid, with total leukocyte counts of less than 5 cells/µL in both areas.²

Pediatric data are more limited. Studies of infants and small children have demonstrated the same types of differences in ventricular and lumbar fluids, taking into account the age-related difference. For example, in children one to four years of age, the mean ventricular protein level is 5 mg/dL versus 15 mg/dL in the lumbar area.³ Consequently, normal cerebrospinal fluids should be location and age specific. Adhering to this, however, is difficult because very limited data may be available for some combinations of age and location.

The fact that a fluid comes from a ventriculoperitoneal shunt adds another complication because a shunt is a foreign body and a foreign body induces a reactive change in the area where it is placed. A study of fluid from patients with ventriculoperitoneal shunts who were evaluated for the possibility of shunt malfunction/infection showed that cell counts were consistently increased compared to normal cerebrospinal fluid. Leukocyte counts ranged from a mean of 43 cells/µL in patients with neither shunt dysfunction nor infection, to 49 cells/µL with an increased eosinophil fraction in patients with shunt malfunction who had no infection, to 2,197 cells/µL with an increased neutrophil fraction in patients with infection.⁴ Thus, cerebrospinal fluid from ventriculoperitoneal shunts does not demonstrate results that correspond to normal lumbar values, but the "normal" range for such a fluid is undefined.

The best the laboratory can do for such a fluid is indicate that low-level pleocytosis can be seen as a result of the presence of a shunt and that significant pleocytosis or an increase in the fraction of eosinophils (greater than five percent) or neutrophils (greater than 10 percent) should alert the clinician to shunt malfunction or infection.

Reference

1. Gerber J, Tumani H, Kolenda H, et al. Lumbar and ventricular CSF protein, leukocytes, and lactate in suspected bacterial CNS infections. Neurology. 1998; 51:1710–1714.

2. Wong M, Schlaggar BL, Buller RS, et al. Cerebrospinal fluid protein concentration in pediatric patients: defining clinically relevant reference ranges. Arch Pediatr Adolesc Med. 2000;154:827–831.

3. Kjeldsberg C, Knight J. Cerebrospinal fluid. In: Body Fluids. 3rd ed. Chicago, Ill.: ASCP Press;1996:65–158.

4. McClinton D, Carraccio C, Englander R. Predictors of ventriculoperitoneal shunt pathology. Pediatr Infect Dis J. 2001:20:593–597.

Robert Novak, MD
Department of Pathology
Children’s Hospital
Medical Center of Akron (Ohio)
Chair, CAP Hematology/Clinical Microscopy Resource Committee