Preserving routine urine specimens, documentation of normal flora, and quantitative body fluid counts. (Tips from the Clinical Experts).

Urine preservation

Q I am looking for an easy way of preserving routine urine specimens that will not be tested within two hours of collection. Presently, in-house specimens in our 300-bed hospital are collected from the nursing units every two hours by a laboratory messenger. Many times, these specimens are almost two hours old when they arrive in the lab. Specimens from our outreach clients are to be refrigerated, but often they stand in the outreach area for an hour or two before reaching the urinalysis section.

I have recommended that all specimens collected by the nursing units or held in the outreach section be placed in Styrofoam coolers containing freezer packs until they are brought to the urinalysis section. The freezer packs could be replaced daily by the lab messenger. Our lab manager is not in favor of this recommendation and has asked me to look for a chemical preservative. The search for a preservative other than refrigeration for a routine urinalysis has not been successful. What do you suggest?

A According to NCCLS Approved Guidelines on Urinalysis and Collection, Transportation, and Preservation of Urine Specimens, "In general, chemical preservatives should be avoided for urinalysis." (1) NCCLS recommends that urinalysis be performed within two hours of collection. If this is not possible, the urine specimen should be refrigerated as soon as possible after collection. The length of time a specimen can be held under refrigeration for adequate preservation has not been agreed upon -- different urine constituents remain unchanged under refrigeration for different lengths of time. Generally, the urinalysis should be completed within six to eight hours of collection. Decomposition of urine begins within 30 minutes; therefore, ideally, urine should be examined within this time. Urine left at room temperature for more than two hours is usually considered unacceptable for examination. Refrigeration is generally adequate for constituents encountered in the routine urinalysis, with the exception of bilirubin and urobilinogen, which are labile to both heat and light. Refrigeration can result in the precipitation of urates or phosphates, which may obscure other pathologic constituents in the microscopic examination of the urine sediment. According to NCCLS, "If the urine is also to be cultured, it should be refrigerated during transit and held refrigerated until cultured." (1)

There are urine preservation tubes commercially available for both urinalysis and culture that are said to preserve urine up to 72 hours without refrigeration. They are generally screw-on cap, leak-proof polypropylene tubes with conical bottoms and skirted freestanding bases, and each contains a preservative tablet such as mercuric oxide. The capacity of the tube is limited -- as little as 10mL. Culture tubes are also flat-bottomed and contain a different preservative such as boric acid-based preservative tablets. These tubes are not interchangeable; specimens that are to be cultured must be placed in the culture tube, and specimens for routine urinalysis, in the tube meant for urinalysis. Preservatives that interfere with any of the chemical tests included in the routine urinalysis are not acceptable. Since the preservation tubes are fairly small, a suitable collection container must be provided to the patient for collection of the sample, and the urine must be well mixed and poured into the preservation tub e -- all introducing a chance of processing error, plus the additional cost of the containers.

Many of the changes that occur in urine stored at room temperature relate to multiplication of bacteria. Preservatives for urinalysis, including refrigeration, generally work by inhibition of bacterial growth. The use of these chemical preservatives result in a specimen that is unsuitable for culture. Changes include an increase in urinary pH as urea is broken down to ammonia. In addition, casts tend to decompose, and red cells undergo hemolysis. Turbidity increases due to the growth of bacteria. The glucose level decreases due to metabolism by bacteria, while nitrates are converted to nitrite through bacteria action.

Other changes include a darkening of the urine color, due to oxidation of colorless chromogens such as the oxidation of urobilinogen to urobilin, or a change in color as bilirubin is oxidized to biliverdin. In alkaline urines of a low specific gravity, cells and casts begin to lyse. Leukocytes are especially subject to lysis as the urine stands at room temperature. If urine specimens contain glucose, the presence of leukocytes will result in decreased levels of glucose due to metabolism by the cells. If ketones are present, the level will decrease as acetone is converted to water and carbon dioxide or the ketone volatilizes.

In summary, refrigeration appears to be the best solution when urinalysis cannot be performed within two hours of collection. Your recommendation that all specimens be placed in coolers containing freezer packs until they are brought to the urinalysis section (assuming it is impossible to refrigerate the specimens at the collection site) seems to be an acceptable alternative to examination within two hours of collection.

--Karen M. Ringsrud, MT(ASCP)

Assistant Professor

Dept. of Laboratory Medicine and Pathology

University of Minnesota Medical School 

Reference

(1.) NCCLS. Urinalysis and Collection, Transportation, and Preservation of Urine Specimens: Approved Guidelines. Villanova, PA: National Committee for Clinical Laboratory Standards; 2001;21(19):GP16-A2.

Normal flora documentation

Q Are we required to document culture results of "no growth" or "normal flora" on the culture worksheet, or can we issue the report without documentation?

A The answer to this question is linked to the need for a culture worksheet. The worksheet, whether it is paper or on a computer, serves two functions. First, the worksheet provides documentation of all work performed on the culture by previous individuals, and second, it provides the information needed to reconstruct the work performed on the culture months or years later.

Therefore, the worksheet should clearly and accurately record all observations, smears, test results, telephone calls to providers, and actions taken at each step of the culture workup as required by the procedure manual. The procedure manual dictates the extent of the required documentation. For example, when the procedure manual states that a bile solubility test is used to differentiate Streptococcus pneumoniae from viridans streptococci in sputum cultures, the results of that test should be recorded on the worksheet. Or, when the procedure requires examination of the culture plate each day for three days, a notation should be made indicating that the necessary examinations have occurred. These notations assist the different individuals who may work on the same culture. Often, the documentation of negative results on the worksheet appears to be a waste of precious time, but it does serve as a record that the culture report is correct based on the work performed according to accepted laboratory procedure.

--David Sewell, Ph.D., ABMM 

Director of Microbiology

Veterans Affairs Medical center

Portland, OR

Quantitative body fluid counts

Q We currently offer cell counts on fluids such as cerebrospinal fluids, paracenteses, synovial fluids, and other fluids. I can see the importance of reporting both WBC and RBC counts on cerebrospinal fluids, but how important is a cell count for the other fluids? Is it acceptable to report cell counts semiquantitatively, e.g., RBC 1+, 2+, etc., for these types of fluids?

A Currently, it is recommended that cell counts on spinal fluids be quantitatively determined, rather than only estimated as 1+, 2+, etc. (1) This is because the definitions of such semiquantitative intervals are not yet standardized; therefore, comparisons with reports from the literature could be problematic. The examination of spinal fluids for evidence of infection, including bacterial vs. viral, or cerebral hemorrhage, is particularly important for patient care. There is a temptation to do the quantitative cell counts on a hematology analyzer, although the manufacturers may not recommend that method. But with the increased accuracy and precision of these counters, I would expect contemporary studies will obviate this objection. Also, the development of cytocentrifugation techniques for spinal fluids, as well as other body fluids, makes this method very attractive for differential counting. There are, however, the necessities of purchasing and maintaining an additional piece of equipment and the purchase of disposable devices for the preparation of microscopic slides.

Quantitative cell counts on other fluids, such as urine, have a considerable benefits. Interestingly, microbiology laboratories are now using accurate estimates of cell counts on urinalysis to decide which urine specimens should be cultured, and significant savings of time and resources have been achieved. As counts on other fluids improve, hopefully similar improvements would be found for other fluid examinations.

Of course, in addition to quantitative counting, the morphology is also important. Synovial fluids occasionally are submitted for examination and identification of any diagnostic crystals as might be seen in gout or other types of arthritis. All of these fluids can conceivably contain malignant cells; therefore, the possibility of such cells must be kept in mind.

--John A. Koepke, M.D.

Professor Emeritus of Pathology

Duke University Medical Center

Durham, NC

Reference

(1.) Gall JJ. Laboratory evaluation of body fluids. In: Stiene-Martin E, Lotspeich-Steininger CA, Koepke JA, eds. Clinical Hematology: Principles, Procedures, Correlations. 2nd ed. Philadelphia: Lippincott; 1997:400-414.

Daniel M. Baer is professor emeritus of laboratory medicine at Oregon Health and Science University in Portland, OR, and a member of MLO's editorial advisory board.