Tips Edited by Daniel M. Baer, MD

42 July 2004 ■ MLO www.mlo-online.com

MLO’s TTTTTips from the Clinical Expertsips from the Clinical Expertsips from the Clinical Expertsips from the Clinical Expertsips from the Clinical Experts department provides practical, up-to-date solutions to readers’ technical and clinical issues from a panel of experts in variousfields. Readers may send questions to Dan Baer by e-mail at

Tips from the clinical expertsEdited by Daniel M. Baer, MD

42 July 2004 ■ MLO www.mlo-online.com

MLO’s TTTTTips from the Clinical Expertsips from the Clinical Expertsips from the Clinical Expertsips from the Clinical Expertsips from the Clinical Experts department provides practical, up-to-date solutions to readers’ technical and clinical issues from a panel of experts in variousfields. Readers may send questions to Dan Baer by e-mail at tips@mlo-online.com.

Reports have shown,predominately in cases of

severe bacteremia or fulminatesepsis, that organisms can be

seen in peripheral bloodsmears prepared from bloodanticoagulated with EDTA.

Pyridium crystals in a urine sediment

QAre there such things asPyridium crystals in a urine

sediment?

ATo my knowledge, crystals ofPyridium have not been described

in urinary sediment. If other crystalsare present in a urine specimen, how-ever, it is quite possible that theycould be stained with the Pyridiumdye. This shows the importance ofshape when identifying crystals in theurine sediment.

Pyridium (Parke-Davis) orphenazopyridine is used as an analge-sic for relief when pain, burning, ur-gency, frequency, and other discom-forts that result from irritation of themucosa of the lower urinary tract.These symptoms may be the result ofinfection, trauma, surgery, endoscopicprocedures, or the passage of stonesor catheters. It is used for symptom-atic relief of pain and not treatment.

Phenazopyridine produces a brightorange- to red-colored urine speci-men. It resembles the color of urinecontaining bilirubin or urobilinogen,although the color is more vivid. Itspresence presents a problem becauseit interferes with several reagent striptests by masking or obscuring thecolor reaction, leading to false-posi-tive results. Various constituents thatmight be present in the urine sedi-ment, such as cells or casts — and atleast, theoretically, crystals — mightalso be stained by the presence ofphenazopyridine.

—Karen M. Ringsrud, MT (ASCP)Assistant Professor

Department of Laboratory Medicineand Pathology

University of Minnesota Medical SchoolMinneapolis, MN

Gram stains from EDTA tubes

QDo you have any referencesthat address the issue of using

specimens that are received in ster-

Answering your questionsile vacutainer tubes with EDTA ad-ditive (liquid or dry) for Gram-stainexaminations? I know that EDTAand other additives can be toxic tobacterial specimens placed in thistype of container and are, there-fore, not suitable for culturing;however, I do not know of a refer-ence that indicates that it is notsuited for holding specimens forGram staining. Will the EDTAcause the bacteria to lyse if it isplaced in this medium? On rareoccasions, I have examined periph-eral smears from EDTA specimensfrom patients that are septic andcontain either intracellular or ex-tracellular cocci or bacilli, so I amaware that at least some bacteriacan be safely transported in thismedium for this purpose.

AThere are several examples ofcases where microorganisms have

been identified in blood smears orbuffy-coat preparations made fromEDTA-containing tubes, using bothroutine hematology stains and/or theGram stain.1-9 I am unaware, however,of a reference that specifically detailsthe use of EDTA as a preservative or a“holding medium” for specimensawaiting a Gram stain.

Reports have shown, predominatelyin cases of severe bacteremia or fulmi-nate sepsis, that organisms can be seenin peripheral blood smears preparedfrom blood anticoagulated withEDTA.1-6, 8 Keep in mind, however, thatthe possibility of microbial contamina-

tion of the anticoagulant tubes, stain-ing reagents, and/or slides could leadto spurious bacterial findings.5, 6

—Susan E. Sharp, PhD, (DABMM)Director of Microbiology

Kaiser Permanente;Associate Professor

Pathology Regional LaboratoryOregon Health and Science University

Portland, OR

References

1. Pedersen G, Schonheyder HC, Nielsen LC.Capnocytophaga canimorsus bacteraemia demon-strated by a positive blood smear. A case report.APMIS. July 1993;101(7):572-574.

2. Rodwell RL, Leslie AL, Tudehope DI. Evaluation ofdirect and buffy coat films of peripheral blood forthe early detection of bacteraemia. Aust PaediatrJ. April 1989;25(2):83-85.

3. Lehmann LS, Seivak JL. Rapid and definitive diag-nosis of infectious diseases using peripheral bloodsmears. J Intensive Care Med. 1992;7:36-47.

4. Fife A, Hill D, Barton C, Burden P. Gram negative sep-ticaemia diagnosed on peripheral blood smear ap-pearances. J Clin Pathol. January 1994;47(1):82-84.

5. Morgan M. Capnocytophaga canimorsus in pe-ripheral blood smears. J Clin Pathol. July 1994;47(7):681-682.

6. Van der Meer W, Verwiel JM, Gidding CE, De MetzM, De Keijzer MH. Bacteria in blood smears: over-whelming sepsis or trivial contamination. ActaHaematol. 2002;107(4):220-223.

7. Graham BS. Detection of bacteremia and fungemia:microscopic examination of peripheral bloodsmears. Infect Control. September 1984; 5(9):448-452.

8. Storm W. Early detection of bacteremia by periph-eral blood smears in critically ill newborns. ActaPaediatr Scand. 1981;70(3):415-416.

9. Selby DM, Gautier G, Luban NL, Campos JM. Over-whelming neonatal septicemia diagnosed uponexamination of peripheral blood smears. ClinPediatr. December 1990; 29(12):706-709.

Refrigerating urine for uric acidanalysis

QFor the past 30 years that I havebeen working here, we have al-

ways asked our clients to refriger-ate (or cool on ice) 24-hour urinecollections. This is primarily to pre-vent offensive odors. For urine uric-acid analyses, we mix the collectionwell, remove a 3-mL aliquot, adjustthe pH to 10 to 11, heat the aliquotto 56°C for 10 minutes, again mix

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well, then centrifuge and analyze. Recently, I read in anNCCLS guideline that 24-hour collections for uric acidshould not be refrigerated. I have tried finding a refer-ence for our collection and processing method but havenot been successful. Can you help me with this? It wouldbe difficult to validate our method without making thechange, but making the change requires notification ofall our many outreach clients.

ASolute concentration, pH, storage time, and tempera-ture all play a role in crystal formation. Precipitation of

urates is often seen in acidic urine at 4°C. Amorphousurates are the most common urate precipitates and typi-cally are of no clinical significance; other urates includeuric acid, acid urates, and sodium urates.1 Uric-acid crys-tals are commonly seen in patients with gout and tumorlysis syndrome. Precipitated urates dissolve after additionof alkali or upon heating.

Massive amounts of amorphous urates can obscure clinicallysignificant formed elements in urine on microscopic examina-tion.2 Such interference is a potential problem in the study of allurinary sediments and, therefore, a consideration that is notexclusive to urine collections for uric-acid quantitation. Ideally,urine should be handled at a temperature similar to body tem-perature and analyzed quickly.3 Despite the risk of urate pre-cipitation, urine should be refrigerated if analysis is not possiblewithin two hours of collection. Refrigeration suppresses multi-plication of bacteria, preserves urinary casts, and slows any shiftsin urinary pH and osmolality.4

If a 24-hour urine specimen is to be used for quantitativeurate determination, NCCLS Document GP13-P does in-deed state that the specimen should not be refrigerated.5

There is no apparent reason, however, that refrigerationwould specifically affect urate determination, as long asprecipitated urates are returned to solution prior to analy-sis. Interestingly, there is no recommendation in NCCLSDocument GP16-A2 that urine specimens should not berefrigerated prior to urate determination.6 �

—Andrew M. Schreiner, MDDepartment of Pathology

Oregon Health and Science UniversityPortland, OR

References

1. Strasinger SK. Urinalysis and Body Fluids, 4th ed. Philadelphia, PA: FA Davis; 2001.2. Brunzel NA. Fundamentals of Urine and Body Fluid Analysis. Philadelphia, PA:

Saunders; 1994.3. Fogazzi GB, Ponticelli C, Ritz E. The Urinary Sediment: An Integrated View. New

York: Oxford University Press; 1999.4. Over, S. The effect of delay in processing on urine particle analysis. Sysmex Jour-

nal International. 2002; 12:9-12.5. The National Committee for Clinical Laboratory Standards (NCCLS): Collection and

preservation of timed urine specimens; proposed guideline. NCCLS DocumentGP13-P. Vol. 7. No. 8; September 1987.

6. The National Committee for Clinical Laboratory Standards (NCCLS): Urinalysis andcollection, transportation, and preservation of urine specimens; approved guide-line-2nd ed. NCCLS Document GP16-A2. Vol. 21. No. 19; November 2001.

Daniel M. Baer is professor emeritus of laboratory medicine at Oregon Healthand Science University in Portland, OR, and a member of MLO’s editorial advi-sory board.

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Tips from the clinical experts