The routine urinalysis is a quick and relatively inexpensive test which can be readily performed in most practices. The results are useful in a variety of situations and are not limited to those directly involving the urinary tract. Routine urinalysis is an essential part of the diagnostic evaluation of sick patients and the results should be interpreted along with the results of a chemistry panel. Ideally, urine should always be collected at the same time as blood for hematology and clinical chemistry and before any treatment (including intravenous fluids) is administered. Complete interpretation of results of chemistry panels cannot be performed without concurrent knowledge of the urinalysis, particularly if there are abnormalities in renal (e.g. urea nitrogen and creatinine) or acid-base parameters on the chemistry panel. Similarly, interpretation of some abnormalities in urine (e.g. glucosuria, ketonuria) is facilitated by concurrent knowledge of chemistry results. The analyzer used at Cornell University is the CLINITEK Advantus Urine Chemistry Analyzer.
When using the urinalysis to evaluate renal function, the test gives us information on the following:
- Tubular function: A urinalysis gives us multiple aspects of tubular function
- Urine specific gravity (USG) tells us about the ability of the loop of Henle to dilute urine and distal tubules to concentrate urine (yes, extrarenal factors do affect the urine specific gravity).
- Dipstick analysis tells us about the ability of proximal tubules to resorb normally filtered low molecular weight substances, including proteins and glucose. Proteinuria that is excessive for the USG and glucosuria without hyperglycemia can reflect proximal tubular disease (yes, other factors affect these as well).
- Sediment examination can help us detect tubular injury via the presence of cellular, granular, waxy casts or other rare casts (hemoglobin, red and white blood cell).
- Glomerular function: This is generally assessed more from serum or plasma biochemical tests, i.e. urea nitrogen, creatinine and SDMA, however proteinuria (particularly albuminuria) reflects glomerular barrier function. In rare cases, hematuria can be glomerular in origin.
- Other conditions
- Hemorrhage via the presence of excess red blood cells in the urine, usually accompanied by a positive heme reaction on the dipstick.
- Infection or inflammation in the urinary tract via the presence of white cells (neutrophils primarily, pyuria), and/or bacteriuria (not explained by contamination)
- Cancer, if tumor cells exfoliate into the urine.
- Crystalluria: These can be normal, part of an overall metabolic condition (e.g. cystinuria) or responsible for an animal’s urinary tract disease if there are uroliths.
Equipment
Required equipment and materials include:
- microscope
- centrifuge
- refractometer
- multiple reagent dipsticks
Urine should be examined as soon as possible after collection, because artifacts will occur in the urine over time (cells lyse, crystals form in vitro). If a delay is anticipated before analysis, the urine should be refrigerated. Refrigerated urine should always be brought to room temperature before testing.
Procedure
Urinalysis consists of the following steps:
- Assessment of visual urine attributes: This includes volume, color and turbidity. It is ideal to standardize the volume of urine from which the urine sediment is prepared. In human medicine, 10 mL of urine is used as the standard volume. This is difficult to accomplish in many animals, particularly small patients, hence we try and standardize the urine volume from which the urinalysis is performed (regardless of the volume received) to 5 mL in the laboratory at Cornell University. However, we often do not receive even this much urine for analysis. Urine volume affects the results of the urine sediment examination, because the semi-quantitative results of the sediment are derived from the standard urine volume and will differ between urine collections of different volumes. Observations of color and turbidity are made on the well-mixed urine specimen.
- Assessment of concentrating ability: This is usually done by measuring specific gravity with a temperature-compensated urine refractometer that is designed for use in veterinary species. The specific gravity should be read on the urine supernatant after centrifugation of the urine and not on uncentrifuged urine (particulate material could scatter light if in suspension). Concentrating ability can also be assessed more accurately by measuring urine osmolality, but this is not usually done as part of the routine urinalysis.
- Dipstick analysis: Dipsticks consist of various pads containing chemical ingredients which provide a color change when a particular analyte is present in urine. This color change is converted to a semi-quantitative result for the analyte in question. In animals, the dipstick is used to give results for pH, protein (mostly albumin), glucose, ketones (primarily acetoacetic acid), bilirubin (the conjugated or direct form) and proteins containing a heme group (a porphyrin ring with iron in its center). There are also dipstick pads for urine specific gravity, nitrate, leukocytes and urobilinogen on commercially available dipsticks, e.g. Multistix®. These are either not accurate in animals or do not provide much additional information in animals and are seldom, if ever, reported.
The dipstick analysis is usually performed on uncentrifuged urine, unless there is marked hematuria (which may affect interpretation of the color changes on the dipstick). In urine with marked hematuria, the interfering erythrocytes can be sedimented by centrifugation and the dipstick analysis can be performed on the supernatant. To perform the analysis, the dipstick can be literally dipped quickly into the urine to immerse the pads or a transfer (plastic) pipette can be used to drop urine individually onto the pads (we use the latter technique when there is too little urine volume to adequately immerse the dipstick). The level of concordance between results of visual dipstick assessment by different observers with the same urine sample in dogs was assessed in one study, which showed that the dip method is more concordant than the drip method (87% versus 80%). However most discordant results were one level different (one higher or lower, e.g. 1+ versus 2+). More variability was evident with pH, which tends to be less accurate anyhow (Boag et al 2019). With either technique, it is important to remove the excess urine – such as by touching the side of the dipstick onto a piece of gauze. It is also important to not allow the urine to spread from one pad to the other as this may interfere with the chemical reactions. When reading the dipstick (i.e. comparing to the color changes on the container), hold the dipstick flat (horizontal) versus upright (vertical) to avoid excess urine from transferring between pads. When using dipsticks, follow manufacturer’s directions on storage and use of the reagents. The package insert also contains useful information about test limitations and interfering substances.
In clinical practice, the degree of color change on the dipstick (e.g. 1+, 2+) is visually (and subjectively) assessed. At Cornell University, we no longer visually examine the Multistix® to determine the dipstick results, rather the CLINITEK Advantus® Urinary Chemistry Analyzer is used, which provides an automated reading of the Multistix® family of urinalysis tests. The machine corrects for the urine color and provides semi-quantitative results for several of the urine results. - Sediment examination: For this examination, the standard volume of urine is centrifuged in a low speed centrifuge (e.g. 250 g for 5 minutes), although other techniques can also be used, such as gravity sedimentation in a microwell plate (Chase et al 2017). The supernatant is decanted (by rapid inversion once) or removed with a plastic pipette and the urine is gently resuspended in a standard volume (0.5 ml) of urine supernatant with a pipette. A drop of the resuspended urine is placed on a slide, coverslipped, and examined under a light microscope using the 10x and 40x objectives (this is frequently called a “wet prep”). Subdued lighting is necessary to increase refractility of the unstained urine elements (lower the condenser and/or close down the substage iris diaphragm). At Cornell University, we do not use stains (e.g. Sedi-stain) to highlight urinary constituents, we only examine unstained urine sediments. We also have a phase contrast microscope which facilitates examination (helps differentiate casts from mucus or bacteria from amorphous particulate material, helps identify hyaline casts).
To examine a urine sediment, we do the following:- Low magnification: Examine the entire coverslip using the 10x objective. At this magnification, casts, large crystals, large infectious agents (parasitic ova), and other constituents (debris, mucus, other contaminants) are semi-quantified.
- High magnification: Specific structures identified at low magnification (e.g. casts) and several random fields are examined using the 40x (high dry) objective. At this higher magnification, cells (leukocytes, erythrocytes, epithelial cells, sperm), small crystals, small infectious agents (bacteria, fungi) and other constituents (fat droplets, debris, other contaminants) are semi-quantified.
