Thursday, 19 March 2015

Urinary tract infections (UTI) 2

Principles of three different automated urine microscopy analysers - Sysmex UF1000i, Menarini sediMAX and Iris iQ200 
The use of automated system in urine analysis reduces the time and cost used in the manual routine analysis. In addition, automated system analyses a high number of samples in a short time (up to 100 samples per hour). There are a few methods available for automated system which includes the Sysmex UF1000i, Menarini sediMAX and Iris iQ200.

The Sysmex UF1000i method uses the urine flow cytometry technology to detect red blood cells, white blood cells, bacteria, epithelial cells and casts in urine. The automated system dilutes 200µL of the urine sample with citrate buffer solution and cationic surfactant with an addition of 40µL of fluorescent dye to complete the mixture. This mixture is then passed through a sheath of flow which is exposed to a laser beam to detect the particles in the stained urine sample with the fluorescent dye staining the nucleic acid in bacteria. The identification and characterisation of particles are based on the fluorescence intensity, forward scatter light and side scatter light intensity which generates electrical signal patterns by photomultiplier conversion which is then measured.

The Menarini sediMAX is an automated microscopy image based sediment analyser used in urine microscopy. The analyser produces a monolayer of urine sediment by centrifugation in a special cuvette. The sediment is analyzed by a bright field microscope and digital camera to capture and categorize 15 particle images based upon size and shape using image processing software. The digital images can be viewed by a trained laboratory staff with zoom capability. The instrument has a high sensitivity (>80%) in the detection of RBC and WBC but has a low sensitivity (approximately 50%) when compared with manual microscopy.

The Iris iQ200 method uses digital imaging and auto particle recognition to classify and quantify urine particles in uncentrifuged urine based upon size and shape. It classifies images such as WBC, RBC, bacteria, casts, yeast and crystals into categories. The digital images generated are collected, approximately 500 frames per sample, then viewed and classified by a trained laboratory staff. This is usually achieved without the use of manual microscopy.
The advantages and disadvantages of the automated urine systems (Sysmex UF1000i, SediMAX and Iris iQ200) are listed below.

Sysmex UF1000i
Uses uncentrifuged urine samples.
Improved bacteria counting and flags for UTI.
Urgent urine samples can be processed by operating the analyser manually.
Throughput of 100 samples per hour.
Cannot differentiate clinically significant pathologic casts cell from benign casts.
Manual microscopy is carried out on flagged results.
This analyser does not require any reagents.
Requires minimal laboratory technician training to interpret images that are similar to manual microscopy.
Results may be available for viewing from the stored digital images
Throughput of 80 samples per hour.

Yeast is mistaken for RBCs.
Epithelial cells are mistaken for WBCs.
Particle loss may occur due to the ultracentrifugation of urine samples.
Iris iQ200
Uses uncentrifuged urine samples.
Accurately quantifies bacteria.
Results may be available for viewing from the stored digital images
Throughput of up to 101 samples per hour.
Crystals are mistaken for RBCs.
There is no sufficient resolution in order to classify all dysmorphic RBC variants which prompt manual microscopy.
Requires considerable laboratory technician training for viewing digital images that are different from manual microscopy.

Detection of biochemical markers and their significance in urine samples
Biochemical markers in urine such as glucose, bilirubin, nitrite, ketone, specific gravity, blood, pH, protein, leukocytes and urobilinogen can be used for UTI. It can be done conveniently as a point of care test or near patients test outside the laboratory. This can be done manually (dipstick) or automated (Siemens Clinitek and Roche Urisys 1100).
A urine dipstick Bayer urinalysis strip such as the Multistix 10 SG is a narrow plastic strip with squares of different colours on it which represents each of the biochemical markers present. The strip is usually dipped in the urine sample to be tested and colour changes noted within 1 minute. The results are then interpreted as shown in the table below.

Table 2: Biochemical markers and its clinical relevance
Biochemical marker
Clinical relevance
Bacteria presence in urine
Leucocyte esterase
WBCs presence in urine
Possible liver disease
Possible liver disease or RBCs breakdown
Acidity of the urine sample and useful in interpretation of microscopic crystals
Product of fat metabolism
Specific gravity
Concentration of urine
Diabetes mellitus
Glomerular disease
RBC trauma or enzyme deficiencies

Automated systems such as Siemens Clinitek analyser is an easy to use chemistry analyser that is used to read Bayer urinalysis strip such as the Multistix 10 SG. The dipped urinalysis strip is inserted into the analyser and result reported automatically.
The Roche Urisys 1100 urine analyser is a semi automated analyser used for in vitro semi quantitative determination of urine analyte. It is a reflectance photometer designed to read and evaluate the results of urine test strips such as Chemstrip 10MD, Chemstrip 7 and Chemstrip 5 OB. The test strip is placed on the sliding tray and a stepping motor moves it under the reading head which remains stationary. The analyser then reads the reference pad followed by each of the pads on the strip. The reading head contains LEDs that emit light at various wavelengths. The LED emits light of a defined wavelength onto the surface of the test pad at an optimum angle. The light hitting the test zone is reflected proportionally to the colour produced on the test pad and is picked up by the detector, a phototransistor positioned directly above the test zone. The phototransistor sends an analog electrical signal to an A/D converter, which changes it to digital form. The microprocessor then converts this digital reading to a relative reflectance value by referring it to a calibration standard. Finally, the system compares the reflectance value with the defined range limits (reflectance values which are programmed into the analyzer for each parameter) and outputs a semi-quantitative result. Each test pad is read photometrically after about 55-65 seconds. In strongly alkaline urine samples, the Urisys 1100 Urine Analyzer automatically corrects the result of the specific gravity test.

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