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What is an abnormal test strip urinary erythrocyte concentration?

Although examination of urine for red blood cells (RBCs) by automated analysis of test strips is a common, precise, and efficient method with a maximum throughput of up to 300 test strips per hour (1), the reference interval for this procedure is uncertain. Most laboratories consider a negative result as normal, based on findings in referred patients and on population studies showing that urinary tract tumors and other serious diseases are found in those with even trace amounts of blood on test strips (2-4). In two population studies, dipsticks were positive for blood in 3% of men of all ages (3) and in 13% of those over 60 years (4). A few patients with blood in their urine were found to have urothelial malignancies, but most had only a trace amount of RBCs on initial testing, with re-testing yielding intermittently positive results (3, 4); intermittently positive results may be attributable in part to variations in the methodology used because visual inspection by technicians and by patients themselves is less sensitive and less precise than semiautomated reflectance readings of urinalysis dipsticks (5).

Our laboratory receives complaints from physicians about the high frequency of hematuria in their patients. We are unaware of previous studies reporting the distribution of RBC concentrations in a healthy population tested by semiautomated reflectance readings. Recent studies using a urine flow cytometer (UF-100) suggest that the number of RBCs in the urine in healthy individuals should be in the range of 10-20 cells/[micro]L with even higher numbers on dipsticks, which also measure lysed cells (6-8). In the present study we analyzed the urine of 1000 men and women of various ages seen consecutively for screening examinations and used automated analysis of the test strips to determine the reference interval.

Midstream urine samples from outpatient clinics arrived and were tested within 4 h of collection at a regional laboratory in northern Israel. Consecutive samples (n = 1000) from occupational medicine clinics that routinely perform urinalysis of asymptomatic workers were tested by use of a Supertron automated analyzer (Roche Diagnostics Ltd.) with Combur-10 S strips (Roche Diagnostics Ltd.). The patients were all actively working without chronic renal failure or symptomatic diseases and had no significant exposure to chemicals or dyes. We have shown previously that the Supertron test for RBCs is precise and stable over a 24-h storage period (1). We tested the accuracy of the assay by adding known numbers of RBCs to urine specimens. A logistic regression model was used to adjust for the presence of urinary glucose, nitrite, or protein and to determine the independent influence of age and sex on the risk of having [greater than or equal to] 50 RBCs/[micro]L on urinalysis.

After excluding those with leukocyturia, glucosuria, proteinuria, or a positive nitrite result, we were left with 580 of the 744 men (78.0%) and 132 of 256 women (51.6%). After these exclusions the distribution of RBC concentrations in urine for males changed little from that for the entire group (results not shown), whereas after the exclusions a lower proportion of females had [greater than or equal to] 50 RBCs/[micro]L [9 of 132 (6.8%) vs 19 of 124 (15.3%); P <0.05]. The optimum reference interval (95% of the individuals studied, after excluding those with other positive findings on dipstick) for RBCs in urine for men <40 years of age included values up to 25 cells/[micro]L, whereas for men [greater than or equal to] 40 years and for females of all ages, the reference interval included values up to 50 cells/[micro]L (Table 1). On logistic regression, after we forced glucosuria (yes/no), nitrite (positive/ negative), leukocyte esterase (yes/no), and proteinuria [greater than or equal to] 300 mg/L (yes/no) into the model, the odds ratio for [greater than or equal to] 50 RBCs/[micro]L in females compared with males was 1.90 (1.02-3.55), the odds ratio for individuals with leukocyte esterase vs those without leukocyte esterase was 2.56 (1.35-4.83), and the odds ratio for individuals [greater than or equal to] 40 years of age compared with younger individuals was 1.93 (1.08-2.45). We found no significant interaction between age and sex.

We added a known number of RBCs (counted on an Advia 120 Hematology System; Bayer) to urine to give a final concentration in the range of 6.25-400 RBCs/[micro]L. The results by dipstick determination were nearly identical to the expected values [expected (dipstick determinations): 0 cells added (12 negative tests by dipstick), 6.25 cells/[micro]L added (0, 10, 10, 0, and 0 cells/[micro]L by dipstick); 12.5 cells/[micro]L added (10, 10, 10, 25, and 10 cells/[micro]L by dipstick); 25 cells/[micro]L added (25, 25, 25, 25, and 25 cells/[micro]L by dipstick); 50 cells/[micro]L added (25, 50, 50, 50, and 50 cells/[micro]L by dipstick)]. Dipstick results were also highly correlated with final concentrations of 100, 200, and 400 cells/[micro]L (results not shown).

The major finding of this study is that the reference interval for RBCs in urine should include positive results. We have also shown that after calibration, dipstick test results are accurate over various known concentrations of RBCs in urine. The reference interval was age dependent, increasing with age, as has been reported previously with a high-power-field methodology (9). Our results can probably be extrapolated to other analyzers that use similar technologies after calibration with known concentrations of RBCs, a necessary step because adequate commercial controls are not available.

Our semiquantitative results are consistent with those obtained with a urine flow cytometer in a small number of apparently healthy children and adults. In young men, we found that the upper limit of the reference interval should be 25 cells/[micro]L RBCs, whereas Lun et al. (6), using the OF-100 urine flow cytometer, reported that the 95th percentile for RBCs in 141 children was 25.9 cells/[micro]L. Regeniter et al. (7) tested 91 healthy individuals (both males and females; age range, 11-63 years) but could not consider subgroups because of the small sample size. They reported 97.5% percentiles of 13.9/[micro]L for RBCs and 15.7/[micro]L for white blood cells; they also reported that the overall mean OF-100 RBC and white blood cell counts were somewhat lower than the results obtained with the semiquantitative stick method, suggesting that lysed cells contributed to the higher observed values. Recently, Marimoto et al. (8) reported median RBC counts on a flow cytometer of ~10 cells/[micro]L in 64 healthy female students 18-20 years of age. The 90th percentile upper limit ranged from 50 to 600, depending on whether the students were menstruating. The 90th percentile range in our females included 50 cells/[micro]L, which is consistent with the results obtained for nonmenstruating students in that study. The automated quantitative urinalysis analyzer (UF-100) cannot be considered a gold standard because the imprecision of the red cell count for this analyzer is high [CV, 18-31% for 5-60 cells/[micro]L (10)]. Furthermore, such testing does not include lysed RBCs.

A major strength of our study is the large number of screening tests done in apparently healthy individuals. Although we did not have complete medical histories available to us, it is unlikely that concomitant diseases biased our results. All participants were actively employed and received medical clearance to work. Chronic diseases are more prevalent in those 50-59 years of age compared with those 40-49 years of age, but the frequency distribution of RBCs in urine was nearly identical in the two groups. After exclusion of those with glucosuria, proteinuria, leukocyturia, or nitrites in the urine, the reference intervals were unchanged. Finally, because the prevalence of microhematuria is not affected by physical exercise in the 24 h preceding urinalysis in asymptomatic young men (11), leisure physical activity probably did not significantly affect our results.

Standard textbooks do not consider semiquantitative or quantitative urinary red cell analysis but instead continue to recommend that a complete workup be done for any patient with a persistent finding of >3 RBCs per high-power field on urinalysis, regardless of the pretest probability of significant urothelial disease (12,13 ). Recent recommendations support a repeat urinalysis after 48 h to confirm positive dipstick findings and a work-up that includes confirmation of the dipstick results by microscopic urinalysis (14,15) and identification of dysmorphic RBCs (14,15). This approach decreases the sensitivity for detecting serious urothelial diseases because of the intermittent nature of hematuria in patients with serious urothelial disease (2-4,16), the known problems with the precision and accuracy of microscopic urinalysis (2,17,18), with intralaboratory CV of 25-50% (8,19), and with the possibility of concomitant dysmorphic RBCs in the urine in the presence of urothelial cancer (20). It has been pointed out that current data are inadequate to support clear-cut recommendations regarding the management of microscopic hematuria (15).

References

(1.) Froom P, Bieganiec B, Ehrenrich Z, Barak M. Stability of common analytes in urine refrigerated for 24 h before automated analysis by test strips. Clin Chem 2000;46:1384-6.

(2.) Messing EM, Young TB, Hunt VB, Emoto SE, Wehbie JM. The significance of asymptomatic microhematuria in men 50 or more years old: findings of a home screening study using urinary dipsticks. J Urol 1987;137:919-25.

(3.) Ritchie CD, Bevan EA, Collier SJ. Importance of occult haematuria found at screening. BMJ 1986;292:681-3.

(4.) Britton JP, Dowell AC, Whelan P. Dipstick haematuria and bladder cancer in men over 60: results of a community study. BMJ 1989;299:1010-2.

(5.) Peele JD, Gadsden RH, Crews R. Semi-automated vs. visual reading of urinalysis dipsticks. Clin Chem 1977;23:2242-6.

(6.) Lun A, Ziebig R, Hammer H, Otting U, Filler G, Sinha P. Reference values for neonates and children for the UF-100 urine flow cytometer [Letter]. Clin Chem 1999;45:1879-80.

(7.) Regeniter A, Haenni V, Risch L, Kochli HP, Colombo JP, Frei R, et al. Urine analysis performed by flow cytometry: reference range determination and comparison to morphological findings, dipstick chemistry and bacterial culture results--a multicenter study. Clin Nephrol 2001;55:384-92.

(8.) Morimoto M, Yanai H, Shukuya K, Chiba H, Kobayashi K, Matsuno K. Effects of midstream collection and the menstrual cycle on urine particles and dipstick urinalysis among healthy females. Clin Chem 2003;49:188-90.

(9.) Froom P, Gross M, Ribak J, Barzilay J, Benbassat J. The effect of age on the prevalence of asymptomatic microscopic hematuria. Am J Clin Pathol 1986;86:656-7.

(10.) Ben-Ezra J, Bork L, McPherson RA. Evaluation of the Sysmex UF-100 automated urinalysis analyzer. Clin Chem 1998;44:92-5.

(11.) Froom P, Gross M, Froom J, Caine Y, Margaliot S, Benbassat J. Factors associated with microhematuria in asymptomatic young men. Clin Chem 1986;32:2013-5.

(12.) Denker BM, Brenner BM. Cardinal manifestations of renal disease. In: Fauci AS, Braunwald E, Isselbacher KJ, Wilson JD, Martin JB, Kasper DL, et al., eds. Harrison's principles of internal medicine. New York: McGraw-Hill, 1998:281.

(13.) Kokko JP. Renal and genitourinary diseases. In: Goldman L, Bennett JC, eds. Cecil textbook of medicine. Philadelphia: WB Saunders Company, 2000:528.

(14.) Grossfeld GD, Litwin MS, Wolf U Jr, Hricak J, Shuler CL, Agerter DC, et al. Evaluation of asymptomatic microscopic hematuria in adults: the Americian Urological Association best practice policy. Part II: patient evaluation, cytology, voided markers, imaging, cystoscopy, nephrology evaluation and follow-up. Urology 2001;57:604-10.

(15.) Cohen RA, Brown RS. Microscopic hematuria. N Engl J Med 2003;348: 2330-8.

(16.) Froom P, Froom J, Ribak J. Asymptomatic microscopic hematuria--is investigation necessary? J Clin Epidemiol 1997;50:1197-200.

(17.) Bee DE, James GP, Paul KL. Hemoglobinuria and hematuria: accuracy and precision of laboratory diagnosis. Clin Chem 1979;25:1696-9.

(18.) Gadeholt H. Quantitative estimation of urinary sediment with special regard to sources of error. BMJ 1964;1:1547-50.

(19.) Fenili D, Pirovano B. The automation of sediment analysis using a new urine flow cytometer (UF-100). Clin Chem Lab Med 1998;36:909-17.

(20.) Offringa M, Benbassat J. The value of urinary red cell shape in the diagnosis of glomerular and post-glomerular haematuria. A meta-analysis. Postgrad Med J 1992;68:648-54.

DOI : 10.1373/clinchem.2003.028589

Paul Froom, * Rachel Etzion, and Mira Barak (Central Laboratory of Haifa and Western Galilee, Clalit Health Services; * address correspondence to this author at: Hematology Laboratories, Central Laboratory of Haifa and Western Galilee, Clalit Health Services, Nesher, Israel; fax 972-4-8209094, e-mail paulfr@clalit.org.il)
Table 1. Number of individuals with various concentrations of RBCs in
screened urine according to age and sex. (a)

 Males, n (%)

 20-29 years 30-39 years 40-49 years
RBCs/ (n = 79) (n = 96) (n = 205)
[micro]L

0 69 (87.3) 73 (76.0) 145 (70.7)
5-10 3 (3.8) 6 (6.3) 23 (11.2)
25 4 (5.1) 15 (15.6) 22 (10.7)
50 0 (0.0) 2 (2.1) 10 (4.9)
150 3 (3.8) 0 (0.0) 5 (2.4)
250 0 (0.0) 0 (0.0) 0 (0.0)

 Males, n (%)
 Females,
 50-59 years 60-64 years (b) n (%)
RBCs/ (n = 169) (n = 45) (n = 132)
[micro]L

0 128 (75.7) 33 (73.3) 99 (75.0)
5-10 18 (10.7) 6 (13.0.3) 10 (7.6)
25 16 (9.5) 4 (8.9) 14 (10.6)
50 4 (2.4) 2 (4.4) 4 (3.0)
150 3 (1.8) 0 (0.0) 5 (3.8)
250 0 (0.0) 0 (0.0) 0 (0.0)

(a) The table includes only individuals without other positive
dipstick findings. The percentages add up to more than 100% because of
rounding off.

(b) Age range, 20-64 years.
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Title Annotation:Technical Briefs
Author:Froom, Paul; Etzion, Rachel; Barak, Mira
Publication:Clinical Chemistry
Article Type:Clinical report
Date:Mar 1, 2004
Words:2300
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