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Urinalysis: Comparison between Microscopic Analysis and a New

Automated Microscopy Image-Based Urine Sediment Instrument

Article  in  Clinical laboratory · April 2014

DOI: 10.7754/Clin.Lab.2013.130725 · Source: PubMed

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Clin. Lab. 2014;60:693-697
©Copyright
SHORT COMMUNICATION

Urinalysis: Comparison between Microscopic Analysis and a

New Automated Microscopy Image-Based Urine Sediment Instrument
PAULA V. BOTTINI, MAYARA H. M. MARTINEZ, CÉLIA R. GARLIPP
Division of Clinical Pathology, University of Campinas/UNICAMP, Campinas, SP, Brazil

SUMMARY

Background: Urinalysis is a high demand procedure, with a large amount of manual labor and poorly standard-
ized. Recently a new walk-away automated urine analyzer has been introduced. The aim of this study was to eval-
uate the performance of UriSed as an alternative to the microscopic analysis of urine samples.
Methods: Four hundred mid-stream urine samples from patients with several clinical conditions were analyzed by
bright field microscopy and by UriSed. The validation protocol included studies of precision, carryover, and com-
parison with the gold standard microscopy.
Results: Our data showed that UriSed is a precise method. Repeatability and reproducibility ranged from 8 to
15%. Carryover was negligible. All the elements showed good agreement between both methods, with an extreme-
ly high correlation between the erythrocyte and leukocyte counts (r > 0.95)
Conclusions: UriSed is a precise and accurate alternative to microscopy that allows a better workflow and may
significantly improve turnaround time.
(Clin. Lab. 2014;60:693-697. DOI: 10.7754/Clin.Lab.2013.130725)

KEY WORDS
urinalysis, microscopic examination, automated sedi-
ment analysis
INTRODUCTION
Urinalysis is a high demand procedure which includes
physical, chemical, and microscopic examinations. Al-
though the analysis of urine sediment provides essential
information on the functional state of the kidneys, it is a
labor intensive and time consuming procedure. Besides
that, it is poorly standardized and features a wide inter-
observer variability as several factors may contribute to
the inaccuracy of this method (e.g., centrifugation, dif-
ferences in interpretation of cellular elements, cylinders,
etc.) [1,2]. Automation seems to be the answer to this
issue [3].
In an attempt to automate the microscopic analysis of
urine, some urine analysis systems have been developed
beginning in the late ‘80s. The great advantage of auto-
mated analysis over microscopic procedure is that the
former reduces the inter-observer variability and in-
creases the accuracy and the productivity. While micro-
scopic analysis usually takes several minutes to be per-
formed automation can release a result in about one
minute [4].
A few years ago, a new walk-away automated urine
sediment analyzer based on the KOVA method with
on-screen review of the images was introduced. This
analyzer is basically the automation of the traditional
manual microscopy [5]. The aim of this study was to
evaluate the performance of this image based automated
sediment analyzer (UriSed, also called sediMAX® in
some countries) as an alternative to the microscopic
analysis of urine samples.
MATERIALS AND METHODS
We analyzed 400 mid-stream urine samples from pa-
tients (262 women and 138 men) with several clinical
conditions. Their ages varied from 2 to 91 years (medi-
an = 41 years).
All samples were analyzed by bright field microscopy
_____________________________________________
Short Communication accepted July 24, 2013

Clin. Lab. 4/2014 693

 P. V. BOTTINI et al.

Table 1. Precision of UriSed.

Intra-assay precision Inter-assay precision

LEVEL 1 LEVEL 2 LEVEL 1 LEVEL 2
element/hpf mean ± SD CV mean ± SD CV mean ± SD CV mean ± SD CV
RBC 25.7 ± 2.0 8% 83.6 ± 7.2 9% 25.5 ± 2.0 8% 95.8 ± 7.7 8%
WBC 11.7 ± 1.2 10% 64.7 ± 6.9 11% 16.1 ± 2.4 15% 79. 9 ± 10.6 13%

RBC - red blood cell, WBC - white blood cell hpf - high power field, SD - standard deviation, CV - coefficient of variation.

Table 2. Diagnostic performance of UriSed.

RBC WBC Hyaline Casts Path Casts Crystals Bacteria

Sensitivity (%) 75 72. 52 54 100.0 69.4
Specificity (%) 99 98 69 82 100.0 85.0
Positive Predictive value (%) 94 94 17 27 100.0 68.8
Negative Predictive value (%) 96 92 92 94 100.0 86.2
Agreement (%) 95.0 91 67 79 100.0 80.1
2
 269.812 242.585 7.195 26.686 397.000 120.247
p < 0.001 < 0.001 < 0.05 < 0.001 < 0.001 < 0.001

RBC - red blood cell, WBC - white blood cell, Path - pathological.

using the KOVA® method with the results expressed in
terms of quantitative elements (erythrocytes-RBC and
leukocytes-WBC; arithmetic mean of 10 fields of 400x)
and qualitative elements (casts, crystals, and bacteria,
considering all fields), and by a fully automated sedi-
ment analyzer (UriSed - 77 Elektronika Kft, Budapest,
Hungary).
UriSed is based on microscopic examination of urine
samples (KOVA method). The analyzer homogenizes,
aspires, and transfers the sample to a special disposable
cuvette. The cuvette is centrifuged and placed in mi-
croscopy position and whole-field images are obtained
and evaluated by an auto image evaluation module
(AIEM). Results were expressed as particle concentra-
tion/hpf (high power field). These high definition im-
ages are displayed on-screen and can be stored for fur-
ther review and for educational purposes [5]. All 15 im-
ages of each sample were reviewed by an experienced
analyst and manually edited, if necessary.
The validation protocol was based on local regulation,
on CLSI documents EP05-A2 [6] and EP15-A2 [7], and
on ICSH guideline [8] and included studies of precision
(repeatability/reproducibility) and carryover.
Repeatability (intra assay precision) was obtained from
the analysis of 20 replicates of two urine samples with
different particle numbers. The inter assay precision (re-
producibility) was performed by analyzing two levels of
control samples (Liquichek Urinalysis Control - Biorad)
on 20 consecutive days.
Carryover effect was assessed by testing a high-level
urine sample consecutively in triplicate (H1, H2 and
H3) followed by a low-level sample (L1, L2 and L3).
Carryover was calculated using the following formula:
carryover (%) = [(L1 - L3)/(H1 - L3)] x 100. Accuracy
of the automated method was evaluated by comparison
with microscopic analysis.
Simple linear regression (least square method) was used
to evaluate the correlation between quantitative analysis
(RBC and WBC) by both methods. The Chi-squared
statistical method was used to determine the agreement
between microscopic analysis and UriSed. RBC and
WBC counts > 5/hpf were classified as abnormal. Qual-
itative elements were compared according to the detec-
tion of them by each method.
RESULTS
Intra assay precision (repeatability) and inter assay pre-
cision (reproducibility) ranged from 8 to 15%, depend-
ing on the considered parameter (Table 1).
As expected, there was no carryover since UriSed uses
single-use cuvette for each sample. The results of high
to low carryover returned negative values: -3.2%
(RBC), -1.8% (WBC) and -1.2% (bacteria).
There was a good correlation between the erythrocyte

694 Clin. Lab. 4/2014

 URINALYSIS: MICROSCOPIC AND AUTOMATED ANALYSIS
Figure 1. Distribution of RBC and WBC counts.
and leukocyte counts by both methods (r > 0.95).
Figure 1 shows the distribution of RBC and WBC
counts by manual and automated analysis.
All the elements showed good agreement between mi-
croscopic analysis and UriSed, as seen in Table 2.
The analyzer easily detected the most common crystals
seen in urine samples. The major causes of misclassifi-
cation are the presence of squamous epithelial cells,
mucus or amorphous crystals, emphasizing the impor-
tance of a recently collected mid-stream urine sample.
DISCUSSION
There is no doubt that microscopy is an important part
of urinalysis and may provide useful information. As it
is a labor intensive and time consuming procedure many
efforts have been made to develop a better, cost-effec-
tive strategy without loss in the diagnostic yield.
The first attempt in reducing the number of urine mi-
croscopy analyses was based on urine dipsticks. This
procedure led to conflicting results as studies showed
that a significant percentage of urine samples with nor-
mal chemistry results on the dipsticks had abnormal fin-
dings on microscopic analysis of sediment [9-11].
Studies have demonstrated significant inter-observer
variation in the identification and interpretation of urine
structures [1,12,13]. Automated urine microscopy is
Clin. Lab. 4/2014
able to classify, quantify, and report urine particles and
it seems to be an efficient alternative to reduce the man-
ual workload and improve inter-observer reliability.
Microscopic examination of urine sediment involves
classification and quantification of particles present in
this fluid and automated devices were developed to fa-
cilitate and standardize this procedure. This goal can be
achieved through three systems with different method-
ologies. One of them (Sysmex UF-100/UF-1000i) is
based on the use of fluorescence flow cytometry to clas-
sify the elements present in urine. Results are displayed
as scattergrams and numerical values are reported [2].
Samples whose results have been flagged need to be re-
viewed by microscopy.
Another analytical system is based on flow cell digital
imaging with automatic particle recognition software
(Iris iQ®200) to identify and classify isolated images
based on the texture, contrast, size, and shape of each
element. The images are displayed to the analyst, al-
lowing edition and reclassification, if necessary [14].
These images are considerably different from those ob-
served in microscopy requiring substantial training be-
fore using the equipment [15].
The third analyzer (UriSed) is based on microscopic
urine sediment analysis with digital images and auto-
matic recognition of particles. UriSed corresponds to
the automation of manual microscopy. Due to its char-
acteristics, UriSed eliminates the need of microscopic
695
 P. V. BOTTINI et al.
review of most of the samples. Images are very similar
to those observed by microscopy and can be easily re-
viewed by the analyst.
It is noteworthy that automated urinalysis, enables bet-
ter workflow and may significantly improve turn-
around time, since its processing time is much faster
than the manual method (80 - 100 samples/hour versus
around 10 samples hour/technologist, respectively).
Our data showed that UriSed is a precise method. Inter-
and intra-assay precision ranged from 8% to 15%. This
rate is much lower than that observed in manual micros-
copy where several studies showed a variation of more
than 50% depending on the particle counts [14-16]. Car-
ryover was negligible, as described by Zaman et al.
[17].
All the elements showed good agreement between mi-
croscopic and automated analysis. Our data were in ac-
cordance with those obtained by Akin et al. [14] and
Zaman et al. [17]. Other authors, using automated urine
analyzers based on flow cytometry (UF-100/UF-1000i)
or digital image with particle recognition (iQ®200) also
observed good correlation with the gold standard mi-
croscopy [16,18,19]. Akin et al. compared the analytical
performance of UriSed with another automated analyzer
(iQ®200) and observed a significant correlation between
them [14].
Appropriate preanalytical procedures are the basic con-
dition for obtaining an adequate sample that allows reli-
able results and a proper clinical interpretation. As
stated before, one of the major causes of misclassifica-
tion was the presence of contaminating elements such as
squamous epithelial cells and mucus. Collecting a mid-
stream sample is the most relevant step to prevent con-
tamination. A recent multicenter study conducted by
Manoni et al. [20] clearly confirmed mid-stream urine
as the most suitable sample for routine urinalysis.
In summary, UriSed is a precise and accurate alternative
to the gold standard microscopy. The implementation of
this automated routine allows a better workflow and
may significantly improve turnaround time. Besides
that, it has the potential to eliminate the need of a mi-
croscopic review of most of the samples as UriSed per-
mits an on-screen review of the images. It is noteworthy
that the use of a system that photographs and stores
high-definition images provides an invaluable training
tool for technologists and medical students.
Acknowledgement:
We thank José Ricardo Lauand, Luciane Franco-Fer-
nandes, and Solange Gomes Lara Cioffi for helping us
during the validation process.
Declaration of Interest:
The authors declare that there are no conflicts of inter-
est.
696
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Paula V. Bottini MD, PhD
examination. Scand J Clin Lab Invest 2011;71(1):30-7. Divisão de Patologia Clínica/HC - UNICAMP
Rua Vital Brasil 251- Cidade
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CEP: 13083-888, Campinas, S.P., Brasil
Tel.: + 55 19 3521-7539
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voided urine collection by using automated analyzers for particle Email: paula@hc.unicamp
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