HANDS-ON COURSE |
BEDSIDE URINARY MICROSCOPY |
GIOVANNI BATTISTA FOGAZZI LECTURES SERIES |
URINARY SEDIMENT: Part 1: Methods |
G.B. Fogazzi, Milan, Italy
|
Dr G.B Fogazzi
Research Laboratory on Urine, Unità Operativa di Nefrologia Fondazione IRCCS, Ospedale Maggiore Policlinico, Mangiagalli e Regina Elena Milan, Italy
|
Slide 1
Slide 2
Slide 3
Prof Fogazzi: This is the outline of the course. After a brief introduction, I will
speak about the main methodological aspects concerning the urinary sediment;
the particles of the urinary sediment of nephrological importance with their
clinical implications; the urinary sediment in the clinical practice and,
finally, I will draw some conclusions.
Slide 4
Slide 5
From
the historical point of view, we know that the urinary sediment was introduced
into clinical practice in the late 1830s in
Slide 6
What
is the situation today? In most instances, urinary sediments are examined in central laboratories far from bedside and
without the correct equipment and knowledge, and with the dream to entrust the
whole task to automated instruments. These are already on the market, one being UF 100, which is based on flow cytometry, and the other iQ200, which is based on images
obtained by a video camera. Last but not least, too often the urinary sediment
examination is neglected even by nephrologists.
Slide 7
In
our unit at the Ospedale Maggiore of
Slide 8
Slide 9
Methodological
aspects are very important for urine sediment. The main methodological aspects
include: a correct urine collection; a standardised method for the handling of
the urine; the use of a proper microscope and the use of a proper report to
describe the findings. All these aspects are described in detail in the
document published by the European Urinalysis Group 5 years ago as a supplement
of the Scandinavian Journal of Clinical and Laboratory Investigation (2000; Vol
60, Suppl 231) as well as in our book (Fogazzi GB, Ponticelli C, Ritz E. The
Urinary Sediment. An Integrated View 2nd Edition.
Slide 10
As
to urine collection, it’s important to give the patient written and simple
instructions. According to the strategy of the single lab, we can ask the
patient to supply the first or the second urine of the morning. In our lab, we
ask for the second urine, since
overnight urine, due to its prolonged permanence in the bladder can favour the
lysis of particles. We suggest the patient to avoid strenuous physical effort
in the hours preceding the test, since this may influence in various ways the
findings (for instance by causing haematuria and/or cylindruria). We advise the
patient to clean the external genitalia in an ordinary way. In this respect, we
do not suggest special procedures
since the more the procedures
suggested are complicated, the less the patient is compliant. In order to avoid
contamination, the male has to uncover the glans and female to spread the labia
of the vagina. For the same reason, we always suggest to collect midstream
urine. It’s important to remember that urine collection during menstruation must
be avoided because of the high probability of blood contamination. It is
also important to use a proper urine container (with a capacity of at least 50
to 100 mL, an opening of at least of
Slide 11
What
about a standardized method for the handling of urine? Why is standardization of
the handling of the urine important? It is important because only with a
standardized method we can obtain quantitative reproducible results. The slide
shows the method that we use in our lab. We ask the patient to supply the
second urine of the morning produced over a period of 2 hours; then, we
centrifuge a 10 mL aliquot of urine for 10 minutes at
Slide 12
And
now the microscope. The microscope must be of good quality, must be equipped
with a low and high magnification, and must, and I want to stress must, be
equipped with phase contrast and polarized light.
Slide 13
This
is the microscope we use in our lab. Why phase contrast?
Slide 14
It’s
very simple to explain. On the right you see bright-field, and on the left you
see phase contrast. You see that with phase contrast the particles are much better seen against the background
than with bright-field, and this without the use of stains! I want to stress
that the European Guidelines strongly recommends the use of phase contrast
microscopy.
Slide 15
Slide 16
And
why polarized light? Polarized light is extremely useful to correctly recognize
the crystals. For example, you see uric acid crystals as seen by phase contrast
microscopy (slide 15), and what
happens when we use polarized light (slide 16). Under polarized light, uric
acid crystals assume a typical polychromatic appearance, which is useful to
identify them.
Slide 17
Polarized
light is also important to correctly identify lipid particles,
Slide 18
which
under polarized light appear as “Maltese crosses” which is, “shining” particles
containing a “black cross” whose arms are regular and symmetrical. This feature
allows the identification of lipid particles, especially when they come with an
atypical appearance.
Slide 19
Now,
the urinary sediment report. This is the report we use in our lab. After the
patient details, we have pH, density (or specific gravity), haemoglobin and
leukocyte esterase as detected by dipstick. Then the particles: erythrocytes
(with their morphological classification; see below), leukocytes, tubular
cells, transitional cells (from the deep and superficial layers of the
uroepithelium), squamous cells, casts, lipids, crystals, bacteria, and yeasts.
We also have a space for a brief conclusive comment. I want to stress the
importance of having in the report the findings obtained by dipstick. Why this?
Slide 20
Let
me give you an example. You see in this slide a sample with a pH of 6.0, a
density of 1.006, a +++
haemoglobin and +++ leukocyte esterase by dipstick. However, by microscopy we
find a low number of erythrocytes and leukocytes, which is in contrast with +++
haemoglobin and +++ leukocytes esterase. Is there any explanation for this
discrepancy? Yes there is. The discrepancy is due to the fact that low density
causes the lysis of erythrocytes and leukocytes, which therefore cannot be seen
by microscopy. In contrast, in other instances we may have negative haemoglobin
and many erythrocytes by microscopy. This may be due, for example, to the
presence of large concentrations of Vitamin C in the urine, which reduces the
sensitivity of dipstick for haemoglobin. Therefore, it is always important to
match the findings obtained by dipstick with those obtained with microscopy and
to try to explain them. Thus, our comment for the sample shown in the slide is:
” Mild erythrocyturia and leucocyturia. Please note the discrepancy between
dipstick for haemoglobin and leukocyte esterase and microscopy. This is
probably due to cell lysis caused by low density.” The final message on this point is that examining the urine
only by dipsticks or only by microscopy exposes to the risk of false results.
This risk is reduced when both methods are used on the same sample.
THE SECOND PART WILL BE PUBLISHED ON SEPTEMBER 28TH