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Technical Briefs |
1 College of Medical Technology, Hokkaido University, North-12, West-5, Sapporo 060-0812, Japan
Departments of
2
Laboratory Medicine and
3 Pediatrics, Hokkaido University School of Medicine, North-14, West-5, Sapporo 060-8648, Japan
4 Department of Laboratory Medicine, Kyorin University School of Medicine, 6-20-2, Shinkawa, Mitaka-shi, Tokyo 181-8611, Japan
aauthor for correspondence: fax 81-11-706-4916, e-mail mie{at}cme.hokudai.ac.jp
For urinalysis, midstream collection is recommended (1)(2)(3). Health-associated reference limits for leukocyte and erythrocyte counts in female urine are important for detecting hematuria, pyuria, and urinary tract infection (3). To understand the effects of urinary collection and the menstrual cycle on urinalysis, we examined first-stream and midstream urine samples from healthy female students with use of an automated dipstick reader and the fully automated urine cell analyzer, UF-100.
Specimens were obtained from 64 healthy female students (age range, 18–20 years) at the College of Medical Technology. All were asymptomatic and had no extant urologic disease. They were instructed to collect only first and midstream urine samples in sterile containers at the same time and not to wipe or spread the labia. The volume of the first urine was measured, and the specimen was analyzed within 2 h. The students provided written, informed consent to participate in the study as well as information about their menstrual cycles. Specimens were classified into four groups according to the number of days after menstruation as follows: menstrual (1–7 days after menstruation; n = 15), follicular (8–15 days after menstruation; n = 21), ovulatory (16–19 days after menstruation; n = 6), and luteal (20–31 days after menstruation; n = 22).
Particles in the urine were analyzed by use of a fully automated urine cell analyzer, UF-100 (Sysmex Corporation). We used the manufacturer-defined review flags. Visual review was required for the following: total count >40 000 x 106/L; conductivity <5 or >38 mS/cm; erythrocyte fluorescence >40 channels and erythrocytes >20 x 106/L; percentage of nonlysed erythrocytes <20% and erythrocytes >15 x 106/L; erythrocytes with forward scatter over 200 channels >2.5 x 106/L; casts >2.5 x 106/L; bacteria >1800 x 106/L; pathologic casts >0.5 x 106/L; small round cells >3.0 x 106/L; and yeast-like cells >10 x 106/L. Dipstick urinalysis was performed with N-Multistix SG-L and the automated urine chemistry reagent-strip analyzer, Clinitek 500 (Bayer Medical Corporation). The strips include reagent pads with which to semi-quantify specific gravity, pH, leukocyte esterase, nitrite, protein, glucose, ketones, urobilinogen, bilirubin, and hemoglobin/myoglobin. The Clinitek 500 measures pad color by reflectance colorimetry, and we determined the light intensity of each pad on a photometer at the appropriate wavelength for each color. We used individual original reflectance signals that were continuous within each range.
The median volume of first-stream urine was 17 (range, 8–70) mL. The particle count in each phase is shown in Fig. 1
. Differences between the first-stream urine and the midstream urine were statistically significant for leukocytes, epithelial cells, and bacterial counts during all phases. Erythrocyte counts between the two types of urine specimen significantly differed in all phases except the ovulatory phase. Cast counts between the two types of urine specimen did not differ significantly at any phase.
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Midstream collection decreased the number of samples showing review flags from 22 (19 small round cells, 2 yeast-like cells, and 1 cast) to 5 (small round cells; McNemar test, P <0.001). The microscopic observation of reviewed samples revealed that most (26 of 27) contained epithelial cells or leukocyte aggregates.
The leukocyte esterase results were significantly lower in the midstream urine than in first-void urine for samples in the following phases: menstrual, median (interquartile range), 8 (5–16.5) vs 0 (0–2.8); follicular, 19 (5–16.5) vs 3 (0–23); and luteal, 42 (12–195) vs 7 (1–22.8); Wilcoxon signed-rank test, P <0.01. No other dipstick urinalysis data differed significantly between the first stream and midstream urine samples.
Epithelial cell and erythrocyte counts in midstream urine specimens changed significantly during the menstrual cycle (Fig. 1
). Epithelial cell counts during the luteal phase were higher than those during the other three phases in midstream urine. Erythrocyte counts during the menstrual phase were higher than those during the other three phases. Other cell counts did not vary significantly with the menstrual cycle.
The values at the 90th percentiles of the results for the midstream urine samples during the menstrual, follicular, ovulatory, and luteal phases, respectively, were as follows:
The detection of epithelial cells is considered evidence that the first part of the voided urine has been collected (3). The first part of the voided urine contains considerable amounts of vaginal secretions (3). In agreement with this view, epithelial cell counts in midstream samples were significantly lower. In addition, our data provided evidence that the numbers of erythrocytes, leukocytes, and bacteria derived from contamination by vaginal secretions are reduced in midstream urine samples.
Epithelial cell counts in midstream urine differed in a phase-dependent manner. During the menstrual cycle, estrogen influences the proliferation and maturation of the vaginal epithelial cell layers, whereas progesterone is associated with shedding of the superficial epithelial cell layers; consequently, the number of epithelial cells in vaginal smears increases during the luteal phase (4)(5). These physiologic changes in vaginal secretions during the menstrual cycle could explain the significant cyclic variation in the number of epithelial cells in urine samples from women.
In conclusion, although the number of participants was limited and an investigation of the same individuals during the four menstrual phases will provide more appropriate and valid information, our study reconfirmed the importance of midstream collection for testing of urine from women by modern analytical methods. In addition, our results showed that the numbers of epithelial cells and erythrocytes in midstream urine vary significantly during the menstrual cycle.
Acknowledgments
We thank Osamu Ota, Tsuyoshi Yamane, and Aki Yanagawa for excellent technical assistance. This study was supported by Grant-in-Aid No. 13771449 for Scientific Research from the Ministry of Education, Science, and Culture, Japan.
References
The following articles in journals at HighWire Press have cited this article:
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  M. Morimoto, H. Yanai, and K. Matsuno Morphologic Changes in Urine Particles during the Menstrual Cycle Clin. Chem., June 1, 2005; 51(6): 1081 - 1082. [Full Text] [PDF]
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P. Froom, R. Etzion, and M. Barak What Is an Abnormal Test Strip Urinary Erythrocyte Concentration? Clin. Chem., March 1, 2004; 50(3): 673 - 675. [Full Text] [PDF]
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 D. Chan, A. Ong, M. Schoenberg, M. S. Parmar, R. A. Cohen, and R. S. Brown Microscopic Hematuria N. Engl. J. Med., September 25, 2003; 349(13): 1292 - 1293. [Full Text] [PDF]
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