Routine Urinalysis–the Microscopic Exam of Urine Sediment

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Routine Urinalysis–the Microscopic Exam of Urine Sediment

Learning Objectives

After studying this chapter, the student should be able to:

  1. 1. Discuss the importance of standardizing the microscopic examination of urine and describe how this standardization is achieved in the clinical laboratory.
  2. 2. Describe microscopic and staining techniques used to enhance visualization of the formed elements in urinary sediment.
  3. 3. Describe the microscopic appearance and clinical significance of erythrocytes and leukocytes in urine and correlate their presence with the physical and chemical examination of urine.
  4. 4. Describe the microscopic characteristics and location of each type of epithelium found in the urinary tract, that is, squamous, transitional, and renal tubular epithelium (proximal, distal, and collecting duct).
  5. 5. Summarize briefly the clinical significance of increased sloughing of the urinary tract epithelium.
  6. 6. Describe the formation, composition, and clinical significance of urinary cast formation.
  7. 7. State the categories into which casts are classified, discuss the clinical circumstances that result in the formation of each cast type, and correlate the presence of casts with the physical and chemical examination of urine.
  8. 8. Describe the development of urinary crystals, including at least three factors that influence their formation.
  9. 9. Describe the characteristic form of each major type of urinary crystal; categorize each crystal type as being found in acid, neutral, or alkaline urine; and discuss the clinical significance of each crystal type.
  10. 10. Identify the following formed elements found in urine sediment, and discuss their clinical significance:

Key Terms⁎ *1

The standardized quantitative microscopic examination of urine sediment made its clinical laboratory debut in 1926. At that time, Thomas Addis developed a procedure to quantify formed elements in a 12-hour overnight urine collection. The purpose of this test, the Addis count, was to follow the progress of renal diseases, particularly acute glomerulonephritis. Increased numbers of red blood cells (RBCs), white blood cells (WBCs), or casts in the urine indicated disease progression. A disease process was indicated when one or more of the following cell changes occurred: The number of RBCs exceeded 500,000; the number of WBCs exceeded 2 million; or the number of casts exceeded 5000. Because the Addis count was time-consuming and chemical methods are currently available to monitor the progression of renal disease, it is no longer routinely performed despite its ability to accurately detect changes in the excretion of urinary formed elements. However, microscopic examination of urine sediment continues to play an important role in the initial diagnosis as well as in monitoring renal disease.

Standardization of Sediment Preparation

Ensuring the accuracy and precision of the urine microscopic examination requires standardization. This demands that established laboratory protocols for manually preparing the urine sediment, including using the same supplies, step sequences, timing intervals, and equipment, are adhered to by all personnel. Box 7.1 lists various factors that must be established and followed to obtain standardization in the microscopic examination. Note that all personnel must follow all testing aspects consistently to ensure comparable urinalysis results.

Commercial Systems

To achieve consistency, several commercial urinalysis systems are available (Table 7.1). Each system seeks to consistently (1) produce the same concentration of urine or sediment volume; (2) present the same volume of sediment for microscopic examination; and (3) control microscopic variables such as the volume of sediment viewed and the optical properties of the slides. All of these systems surpass the outdated practice of using a drop of urine on a glass slide and covering it with a coverslip. In addition, commercial slides are cost competitive, easy to adapt to, and necessary to ensure reproducible and accurate results.

Table 7.1

Comparison of Selected Standardized Urinalysis Systems
Features Count-10 System (Myers-Stevens Group) KOVA System (Hycor Scientific) UriSystem Features (Fisher HealthCare)
Initial volume of urine used 12 mL 12 mL 12 mL
Final urine volume with sediment 0.8 mL 1.0 mL 0.4 mL
Sediment concentration 15:1 12:1 30:1
Volume of sediment used 6 µL 6 µL 14 µL
Area for viewing 36 mm2 32 mm2 90 mm2
Number of 100× fieldsa 11 10 28
Number of 400× fieldsa 183 163 459
Coverslip type Acrylic Acrylic Acrylic
Number of specimens per slide 10 4, 10 10


aCalculated using a “field of view” diameter for high power (×400) of 0.5 mm and for low power (×100) of 2 mm. The number of fields possible is equal to the area for viewing divided by the area per low- or high-power field. Note that the field of view diameter is determined by the lens systems of the microscope.

Commercial systems feature disposable plastic centrifuge tubes with gradations for consistent urine volume measurement (Fig. 7.1). The tubes are clear, allowing for assessment of urine color and clarity, and conical, which facilitates sediment concentration during centrifugation. The centrifuge tubes of each commercial system are unique. The UriSystem tube (Fisher HealthCare, Houston, TX) is designed such that after centrifugation, it can be decanted with a quick, smooth motion and consistently retains 0.4 mL of urine for sediment resuspension. The KOVA System (Hycor Biomedical, Garden Grove, CA) uses a specially designed pipette (KOVA Petter) that snuggly fits the diameter and shape of the tube to retain 1 mL of urine during decanting. The Count-10 System (V-Tech, Inc., Lake Mary, FL) offers several options to retain 0.8 mL for sediment resuspension. Each commercial system provides tight-fitting plastic caps for the tubes to prevent spillage and aerosol formation during centrifugation.

A laboratory need not purchase all aspects of a commercial system to obtain a standardized urine sediment for microscopic analysis. In fact, laboratories have considerable flexibility and can blend the systems. For example, a laboratory could choose to use KOVA System tubes to prepare the urine sediment but UriSystem slides or the RS2005 Urine Sediment Workstation (DiaSys Ltd., New York, NY), a semiautomated slide system, to view the sediment. Regardless of the system or combination of products used when preparing and performing the microscopic examination of urine sediment, the imperative is that all personnel adhere to established protocols to ensure that accurate and reproducible results are obtained.

Specimen Volume

A concentrated urine sediment is usually prepared for the microscopic examination. To ensure that a representative sampling of the formed elements in the portion is removed, the urine specimen must be well mixed. The concentrated sediment can be prepared using a variety of initial urine volumes. Frequently, the initial volume of urine is 12 mL with a 12 to 1 concentration (12:1) of sediment prepared for microscopic viewing. However, initial urine volumes ranging from 3 to 15 mL can be used.

Testing using alternate volumes when insufficient specimen is available can be achieved in two ways. One approach is to prepare the same sediment concentration by decreasing the volume of supernatant urine used to resuspend the sediment. For example, suppose the procedure details a 12:1 concentrated sediment (i.e., 12 mL initial urine with the sediment resuspended in 1 mL supernatant urine). When only 6 mL initial urine is available, the procedure is followed but the urine sediment would be resuspended in 0.50 mL—one half the volume used with 12 mL initial urine—to obtain a 12:1 concentration.

A second approach is to reduce the initial volume of urine used and multiply all numeric counts by the appropriate factor. For example, assume as previously that the procedure details a 12:1 concentration of the sediment. When only 6 mL is used, the procedure is followed and all numeric counts from the microscopic exam of the sediment are multiplied by two.

When insufficient urine is available for testing (e.g., <3 mL), well-mixed urine may be examined unconcentrated and an appropriate comment appended to the report. This comment will enable healthcare providers to evaluate the results appropriately. Note that the laboratory reference ranges for a routine microscopic exam will not apply to this unconcentrated sample.

Whenever the actual volume used to prepare the sediment for the microscopic examination is less than that routinely required, a notation should accompany the specimen report. The decision to accept specimens with volumes less than 12 mL for urinalysis, as well as the protocol used for testing, is determined by each individual laboratory.


After well-mixed urine is poured into a centrifuge tube, it is covered and centrifuged at 400 to 450 g for 5 minutes. This centrifugation speed allows for optimal sediment concentration without damaging fragile formed elements such as cellular casts. All personnel must adhere to this 5-minute centrifugation time with all specimens to ensure uniformity. Note that the speed is given in relative centrifugal force (RCF, g) because this term is not dependent on the centrifuge used. In contrast, the speed in revolutions per minute (RPM) required to obtain 400 to 450 g varies with each centrifuge and is directly dependent on the rotor size. For example, one centrifuge may obtain 450 × g at 1200 RPM, whereas another may require 1500 RPM to obtain this same g-force. The RPM necessary to achieve 400 to 450 g can be determined from a nomogram or by using Equation 7.1.

RCF (g)=1.118×105×radius (cm)×RPM2 Equation 7.1

image Equation 7.1

In this equation, the radius in centimeters refers to the distance from the center of the rotor to the outermost point of the cup, tube, or trunnion when the rotor is in motion (Fig. 7.2).

It is important that the centrifuge brake is not used because this will cause the sediment to resuspend, resulting in erroneously decreased numbers of formed elements in the concentrated sediment. In many laboratories, multiple personnel use centrifuges to perform numerous and varied procedures. If all centrifuge settings, including the brake, are not checked before use, the resultant urine sediments can show dramatic variations in their formed elements because of processing differences in speed, time, or braking. Using control materials for the microscopic examination or performing interlaboratory duplicate testing is valuable in its ability to detect these important changes in sediment preparation.

Sediment Concentration

After centrifugation, the covered urine specimens should be carefully removed and the sediments concentrated using the established protocol. Standardized commercial systems accomplish this task through consistent retention of a specific volume of urine. Note that different brands of centrifuge tubes and pipettes should not be intermixed. This can cause variation in the volume of urine retained, which will change the concentration of the sediment. Table 7.1 shows how commercial systems vary in the sediment concentration produced, ranging from a 12:1 to a 30:1 concentration. Manual techniques traditionally strive toward a 12:1 concentration, in which 12 mL initial urine was used; therefore supernatant urine is removed by decanting or using a pipette until 1 mL of urine is retained. Then, a pipette is used to gently resuspend the sediment. Note that too vigorous agitation of the sediment can cause fragile and brittle formed elements, such as RBC casts and waxy casts, to break into pieces.

Volume of Sediment Viewed

A standardized slide should be used for the microscopic examination of urine sediment to ensure that the same volume of sediment is presented for viewing each time. Commercial standardized slides are made of molded plastic and have a built-in coverslip or provide a glass coverslip for use (Fig. 7.3). With a disposable transfer pipette, urine sediment is presented to a chamber, which fills by capillary action. This technique facilitates uniform distribution of the formed elements throughout the viewing area of the slide.

Glass microscope slides and coverslips are not recommended because they do not yield standardized, reproducible results.1 If glass slides are used, the laboratorian should always pipette an exact amount (e.g., 15 μL) of the resuspended sediment onto the glass slide using a calibrated pipette. The volume of sediment dispensed is determined by each laboratory and depends on the size of the coverslip used. The urine sediment volume must fill the entire area beneath the coverslip without excess. Bubbles and uneven distribution of the sediment components can result when the coverslip is applied (e.g., heavier components such as casts are pushed or concentrate near the coverslip edges). If the microscopic examination reveals that the distribution of formed elements is uneven, a new suspension of the sediment should be prepared for viewing. Because all commercial systems have proved superior to the “drop on a slide” method, this technique should not be used for the microscopic examination of urine.2

Reporting Formats

In a manual microscopic examination, urine components are assessed or enumerated using at least 10 low-power (lpf) or 10 high-power fields (hpf). The quantity of some components (e.g., mucus, crystals, bacteria) is qualitatively assessed per field of view (FOV) in descriptive or numeric terms. Table 7.2 lists commonly used terms and typical descriptions. Each laboratory determines which terms are used, as well as the definition for each term. Other sediment components (RBCs, WBC, casts) are enumerated as a range of formed elements present (e.g., 0–2, 3–5, 6–10). Note that although a component may be reported using low-power magnification, high-power magnification may be needed to specifically identify or categorize it; for example, to identify the cell type in a cellular cast. In this case the cells were determined to be RBCs, and the quantity of cellular casts present is reported as the average number viewed using low power (e.g., 3–5 RBC casts/lpf).

Table 7.2

Qualitative Terms and Descriptions for Fields of View
Term Term Description
Rare 1 + Present but hard to find
Few 1 + One (or more) present in almost every FOV
Moderate 2 + Easy to find; number present in FOV varies; “more than few, less than many”
Many 3 + Prominent; large number present in all FOVs
Packed 4 + FOV is crowded by or overwhelmed with the elements

FOV, Field of view.

When a microscopic examination is performed, the volume of sediment viewed in each microscopic FOV is determined by two factors: the optical lenses of the microscope and the standardized slide system used. The ocular field number of the microscope and the objective lens determine the area of the FOV (see Chapter 18). The larger the FOV, the greater is the number of components that may be visible. To obtain reproducible results when manual microscopic examinations are performed, the same microscope must be used, or when multiple microscopes are available, the diameters of their FOVs (i.e., ocular field numbers) must be identical.

These viewing factors and sediment preparation protocols account for the differences observed in reference ranges for microscopic formed elements. They also prevent comparison of the microscopic results obtained in laboratories using different microscopes and commercial slides. However, if each laboratory would relate sediment elements as the “number present per volume of urine” instead of per low- or high-power field, interlaboratory result comparisons would be possible and comparisons between manual and automated microscopy systems (e.g., iQ200 [Iris Urinalysis-Beckman Coulter, Inc., Brea, CA]; UF-1000i [bioMerieux Inc., Durham, NC]) would be facilitated.

To convert the number of formed elements observed per low- or high-power field to the number present per milliliter of urine tested, a few calculations are necessary (Box 7.2). First, the area of the FOV for the low- and high-power fields must be determined. This calculation uses the diameter for the FOV, which is determined by the ocular field number of the microscope and the formula for the area of a circle (area = πr2). Because a standardized commercial microscope slide provides the same volume of sediment in a known viewing area (see Table 7.1) and the area viewed in each microscopic field is known, the “field conversion factors” remain constant. Once the field conversion factors for a particular microscope and the standardized microscope slide system used have been established, determining the number of formed elements per milliliter of urine requires a single multiplication step. Box 7.2 outlines these calculations and includes an example.

Box 7.2

Conversion of the Number of Formed Elements Present in a Microscopic Field to the Number of Formed Elements Present in a Volume of Urine

Calculate the maximum number of low-power and high-power fields possible using your microscope and the standardized microscope slides (see manufacturer’s information or Table 7.1) in use as follows:

Total  coversliparea  for  viewingArea  per  HPF(or LPF)=Number of view  fields  possible


For example, using a KOVA slide and the microscopic lenses given in step 1,

32  mm20.196  mm2=163  fields  of  view  possible  using  high  power


Calculate the field conversion factor, which is the number of microscope fields per milliliter of urine tested, as follows:

Number of view fields  possibleVolume  of  sedimentviewed (mL)×Concentrationfactor=Number of  view  fields1  mL  of  urine  tested


For example, using the KOVA system specimen preparation and a KOVA slide,

163 HPFs  possible0.006 mL sediment  ×12=2260 HPFsmL urine=Field  Conversion  Factor


Convert the number of formed elements observed per HPF (or LPF) to the number present per milliliter of urine by multiplying the number observed per view field by the appropriate field conversion factor.

For example, 2 red blood cells (RBCs) are observed per HPF. Therefore,

2 RBCsHPF×2260 HPFsm L urine=4520 RBCsmL urine


Enhancing Urine Sediment Visualization

When using brightfield microscopy, it can be difficult to see urine sediment components (e.g., mucus, hyaline casts) that have a similar refractive index to that of urine. Because their refractive indexes are similar, there is insufficient contrast to enable optimal viewing. Staining changes the refractive index of formed elements and increases their visibility. Another approach is to change the type of microscopy, which can also facilitate visualization of low-refractility components or can be used to confirm the identity of suspected substances such as fat. Hyaline casts, mucus threads, and bacteria are difficult to see under brightfield microscopy; the use of stains or phase microscopy enhances their visualization. These techniques facilitate observation of the fine detail necessary for specific identification (e.g., distinguishing a WBC from a renal tubular cell). They also help to differentiate look-alike entities, such as monohydrate calcium oxalate crystals, which can resemble RBCs, and can be used to distinguish between mucus threads and hyaline casts. Table 7.3 summarizes the visualization techniques discussed in this chapter.

Table 7.3

Visualization Techniques to Aid in the Microscopic Examination of Urine Sediment
Technique Features
Staining Techniques
0.5% toluidine blue
2% acetic acid
Fat stains: Sudan III, il red O
Gram stain
Prussian blue reaction
Hansel stain
Microscopic Techniques
Phase contrast microscopy
Interference contrast microscopy
Polarizing microscopy

  Casts, mucus Fibers (clothing, diapers), plastic fragments  



RBC, Red blood cell; WBC, white blood cell.

Staining Techniques

Supravital Stains

Numerous stains have been used to enhance the visualization of urine sediment. Each laboratory should have a stain available because stains are inexpensive and can significantly assist in the identification of some urine sediment components. The most commonly used stain is a supravital stain consisting of crystal-violet and safranin, also known as the Sternheimer-Malbin stain (Fig. 7.4). This stain enhances formed element identification by enabling more detailed viewing of internal structures, particularly of WBCs, epithelial cells, and casts. Other formed elements (e.g., RBCs, mucus) stain characteristically, and their descriptions are noted on the package inserts provided with commercially prepared stains. Stabilized modifications of Sternheimer-Malbin stain are available commercially (e.g., Sedi-Stain, Becton, Dickinson and Company, Franklin Lakes, NJ), or it can be prepared by the laboratory if desired.3 One disadvantage of its use is that in strongly alkaline urines, this stain can precipitate, which obstructs the visualization of sediment components.

Another good supravital stain for urine sediment is a 0.5% solution of toluidine blue (Figs. 7.5 and 7.6). The stain is a metachromatic dye that stains various cell components differently; hence, the differentiation between the nucleus and the cytoplasm becomes more apparent. The toluidine blue stain enhances the specific identification of cells and aids in distinguishing cells of similar size, such as leukocytes from renal collecting duct cells.

Acetic Acid

Although acetic acid is not actually a stain, it can be helpful in identifying WBCs. WBCs can appear small, especially in hypertonic urine, with their nuclei and granulation not readily apparent. By adding 1 to 2 drops of a 2% solution of acetic acid to a few drops of urine sediment, the nuclear pattern of WBCs and epithelial cells is accentuated, whereas RBCs are lysed.

Fat or Lipid Stains

Sudan III or oil red O is often used to confirm the presence of neutral fat or triglyceride suspected during the microscopic examination (Fig. 7.7). These lipids stain orange or red and may be found (1) free floating as droplets or globules; (2) within renal cells or macrophages, aptly termed oval fat bodies; or (3) within the matrix of casts as droplets or oval fat bodies. An important note is that only neutral fats (e.g., triglycerides) stain. In contrast, cholesterol and cholesterol esters do not stain and must be confirmed by polarizing microscopy. The distinction between triglyceride and cholesterol is primarily academic because the implications for renal disease are the same regardless of the identity of the fat. In other words, changes have occurred in the glomeruli such that triglycerides and cholesterol from the bloodstream are now passing the glomerular filtration barriers with the plasma ultrafiltrate. The urinalysis laboratory can use a fat stain or polarizing microscopy to confirm the presence of fat; the confirmation method selected is usually determined by cost, personnel preference, and convenience.

Gram Stain

Although Gram stain is used primarily in the microbiology laboratory, it may at times be used in the urinalysis laboratory. Gram stain provides a means of positively identifying bacteria in the urine and differentiating them as Gram negative or Gram positive (Fig. 7.8). To perform a Gram stain, a dry preparation of the urine sediment is made on a microscope slide by smearing and air drying or by cytocentrifugation. As in the microbiology laboratory, the slide is heat fixed and then stained. Gram-negative bacteria appear pink, whereas Gram-positive bacteria appear dark purple. Because these slides can be viewed using a high-power oil immersion (×100) objective, additional characterization of the bacteria (e.g., cocci, rods) could be made, but this is rarely done by the urinalysis laboratory.

Prussian Blue Reaction

To facilitate the visualization of hemosiderin, free floating or in epithelial cells and casts, the Prussian blue reaction, also known as the Rous test, is used. First described by Rous in 1918 to identify urinary siderosis, the Prussian blue reaction stains the iron of hemosiderin granules a characteristic blue.4 See “Hemosiderin” later in this chapter for more discussion of this reaction and its use.

Hansel Stain

Hansel stain (methylene blue and eosin-Y in methanol) is used in the urinalysis laboratory specifically to identify eosinophils in the urine (Fig. 7.9). Whereas Wright’s stain or Giemsa stain also distinguishes eosinophils, Hansel stain is preferred.5 Urine eosinophils can be present in a variety of renal or urinary tract disorders, such as urinary tract infections (UTIs), acute tubular necrosis, glomerulonephritis, and acute interstitial nephritis (AIN).

Microscopy Techniques

Identification of urine sediment components is dependent on (1) the ability of the microscopist and (2) the microscope used to perform the analysis. In the United States brightfield microscopy predominates despite its inherent difficulty in detecting and identifying low-refractile entities, such as hyaline casts, ghost RBCs, and bacteria. Therefore phase contrast microscopy and the availability of supravital stains are strongly recommended in the urinalysis testing area.6 Even the most adept microscopists are restricted in their ability to identify entities when limited by inadequate equipment and supplies. A brief overview of microscopy techniques used in urinalysis testing is introduced here. See Chapter 18 for a detailed discussion of the microscope, the role of each component part, and steps for proper adjustment, as well as principles, advantages, and applications for various types of microscopy.

Phase Contrast Microscopy

Phase contrast microscopy is the preferred technique for microscopic examination of urine sediment because it enables (1) evaluation of RBC morphology and (2) detailed visualization and identification of difficult-to-view (translucent or low-refractile) formed elements such as hyaline casts, RBC ghost cells, and bacteria (Fig. 7.10). An added advantage is that microscopic examinations are generally faster to perform because of the enhanced visualization. See Chapter 18 for a detailed discussion of phase contrast microscopy and how variations in the refractive index of formed elements are converted into variations in contrast, thereby revealing low-refractile components.

Polarizing Microscopy

In the urinalysis laboratory, polarizing microscopy is often used to confirm the presence of fat, specifically cholesterol. Cholesterol droplets are birefringent (i.e., they refract light in two directions) and, similar to their counterpart triglycerides, they can be found as free-floating droplets or in cells (oval fat bodies) and casts. In droplet form—within cells, free floating, or in casts—cholesterol produces a characteristic Maltese cross pattern with polarized light (Fig. 7.11A). These droplets appear as orbs against a black background divided into four quadrants forming a bright Maltese-style cross. When a first-order red compensator plate is used, the background becomes red to violet and opposing quadrants in the orbs are yellow or blue, depending on their orientation to the light (Fig. 7.11B). Note that starch granules and some drug crystals show a similar pattern, which is called a pseudo-Maltese cross because the four quadrants produced are variable in size (see Chapter 18, Table 18.1). Other neutral fats, such as fatty acids and triglycerides, cannot be identified using polarizing microscopy because they are not optically active—light passes through them unchanged. For triglyceride or neutral fat identification, see the section “Fat or Lipid Stains” earlier in this chapter.

Polarizing microscopy can also assist in differentiating urine sediment components that may look alike (see Table 7.3). RBCs can be distinguished from monohydrate calcium oxalate crystals, casts or mucus from fibers, and amorphous material from coccoid bacteria. See Chapter 18 for additional information, as well as a procedure for converting a brightfield scope for polarizing microscopy (Box 18.3).

Interference Contrast Microscopy

Chapter 18 discusses two types of interference microscopy. Differential interference contrast (Nomarski) microscopy and modulation contrast (Hoffman) microscopy provide detailed three-dimensional images of high contrast and resolution (Fig. 7.12). Although their use is suited ideally for microscopic examination of the formed elements found in urine sediment, the increased cost often cannot be justified by the traditional urinalysis laboratory. With experience, however, these microscopic techniques are easy to use and less time-consuming than brightfield microscopy because of the enhanced imaging. In addition, once a brightfield microscope has been modified for modulation contrast microscopy, it can easily be used for brightfield, polarizing, and other techniques by simply removing the specialized slit aperture from the light path.

Cytocentrifugation and Cytodiagnostic Urinalysis


Cytocentrifugation is a technique used to produce permanent microscope slides of urine sediment and body fluids (see Chapter 17). Because a monolayer of sediment components is desired, an initial microscopic examination is required to determine the amount or volume of urine sediment to use when preparing the slide. After this step, the appropriate amount of concentrated urine sediment is added to a specially designed cartridge fitted with a microscope slide that is placed in a cytocentrifuge (e.g., Shandon Cytospin, Thermo Shandon, Pittsburgh, PA). After cytocentrifugation, a dry circular monolayer of sediment components remains on the slide. The slide is fixed permanently using an appropriate fixative and is stained. For cytologic studies, Papanicolaou’s stain is preferred; however, if Papanicolaou’s stain is not available, or if time is a factor, Wright’s stain can be used. The end result is a monolayer of the urine sediment components with their structural details greatly enhanced by staining. This enables the quantitation and differentiation of WBCs and epithelial cells in the urine sediment. If desired, these slides can also be viewed using high-power oil immersion objectives and can be retained permanently in the laboratory for later reference or review.

Cytodiagnostic Urinalysis

In 1926, Thomas Addis established the value of identifying increased numbers of urine cellular elements as evidence of disease progression. Today, the ability to perform urine differential cell counts enables identification of and discrimination between renal disease and urinary tract disorders. Although a cytodiagnostic urinalysis should not be performed on all urine specimens, it can play an important role in the early detection of renal allograft rejection and in the differential diagnosis of renal disease. Cytodiagnostic urinalysis involves making a 10:1 concentration of a first morning urine specimen, followed by cytocentrifugation of the urine sediment and Papanicolaou’s staining.7 Although cytodiagnostic urinalysis requires more time to perform, it is uniquely valuable in the identification of blood cell types, cellular fragments, epithelial cells (atypical and neoplastic), cellular inclusions (viral and nonviral), and cellular casts.

Formed Elements in Urine Sediment

A wide range of formed elements can be encountered in the microscopic examination of urine sediment. These formed components can originate from throughout the urinary tract—from the glomerulus to the urethra—or can result from contamination (e.g., menstrual blood, spermatozoa, fibers, starch granules). Many components, such as blood cells and epithelial cells, are cellular; others are chemical precipitates, such as the variety of crystalline and amorphous material that can be present in the sediment. Casts—cylindrical bodies with a glycoprotein matrix—form in the lumen of the renal tubules and are flushed out with the urine. Opportunistic microorganisms such as bacteria, yeast, and trichomonads can also be encountered in urine sediment. Not all of these formed elements indicate an abnormal or pathologic process. However, the presence of large numbers of “abnormal” components is diagnostically significant.

Identifying and enumerating the components found in urine sediment provide a means of monitoring disease progression or resolution. Determining the point at which the amount of each element present indicates a pathologic process requires familiarity with the expected normal or reference interval for each component (Table 7.4). (See Appendix C for reference intervals of all parameters in a complete urinalysis.) Normally, a few RBCs, WBCs, epithelial cells, and hyaline casts are observed in the urine sediment from normal, healthy individuals. Their actual number varies depending on the sediment preparation protocol and the standardized slide system used for the microscopic examination.8 Because changes occur in unpreserved urine, factors such as the type of urine collection and how the specimen has been stored also affect the formed elements observed during microscopic examination.

Table 7.4

Reference Intervals for Microscopic Examinationa
Component Number Magnification
Red blood cells 0–3 Per HPF
White blood cells 0–8 Per HPF
Casts 0–2 hyaline (or finely granularb) Per LPF
Epithelial cells:
 Squamous Few Per LPF
 Transitional Few Per HPF
 Renal Few (0–1) Per HPF
Bacteria and yeast Negative Per HPF
Abnormal crystals None Per LPF


HPF, High-power field (×400); LPF, low-power field (×100).

aUsing the UriSystem. Values vary with concentration of urine sediment, microscope slide technique, and microscope optical properties. See Appendix C for reference intervals for a “complete” urinalysis.

bAfter physical exercise, cast numbers increase and include finely granular casts (1991, Haber).

This section discusses in detail the variety of formed elements possible in urine sediment and presents the origin of each component and its clinical significance, possible variations in shape and composition, and techniques used to facilitate differential identification. A wide range of additional images of urine sediment components can be found in the Urine Sediment Image Gallery at the end of this chapter.

Blood Cells

Red Blood Cells (Erythrocytes)

The name erythrocyte is derived from the Greek word erythros, meaning “red,” and the suffix –cyte, meaning “cell.” Hence these cells are more frequently called red blood cells (RBCs), and this term will be used predominantly throughout this text. RBCs were one of the first cells recognized and described after the discovery of the microscope.

Microscopic Appearance

Because of their small size—approximately 8 μm in diameter and 3 μm in depth—RBCs in urine are viewed and enumerated using high-power magnification. RBCs have no nucleus; they normally appear as smooth biconcave disks, and they are moderately refractile. When suspended in urine sediment, RBCs can be viewed from any angle. When viewed from the side, they have an hourglass shape; when viewed from above, they appear as disks with a central pallor (Fig. 7.13). The size or diameter of RBCs is affected by urine concentration (i.e., osmolality, specific gravity). In hypertonic urine, their diameter can be as small as ~ 3 μm and in hypotonic urine as large as 11.8 μm.9

Dysmorphic or distorted forms of RBCs can also be present in urine (Fig. 7.14). At times, these forms are present with normal RBCs in the urine of healthy individuals. Some dysmorphic forms occur because of the urine’s concentration (i.e., osmolality).10 The most common dysmorphic form is crenated erythrocytes (i.e., echinocytes or burr cells). When RBCs are present in hypertonic urine (osmolality >500 mOsm/L), they become smaller as intracellular water is lost by osmosis, which causes them to become crenated. As they crenate, erythrocytes lose their biconcave disk shape and become spheres covered with evenly spaced spicules or crenations. Because of these reversible membrane changes, the surface of crenated cells appears rough or sometimes grainy, depending on the microscope adjustments, compared with normal erythrocytes. In hypotonic urine (osmolality <180 mOsm/L), erythrocytes swell and will eventually release their hemoglobin to become “ghost” cells, which are cells with intact cell membranes but no hemoglobin. These empty cells, outlined by their membranes, appear as colorless empty circles. Because their hemoglobin has been lost, ghost cells are difficult to see using brightfield microscopy; however, they are readily visible with phase contrast or interference contrast microscopy (Fig. 7.14). Note that alkaline urine promotes RBC lysis and disintegration, which results in ghost cells and erythrocyte remnants.

A variety of dysmorphic erythrocyte forms can be present in a single urine sediment.10 These forms include acanthocytes, schizocytes, stomatocytes, target cells, and teardrop cells (Table 7.5). Some of these forms are reversible and induced by the physical characteristics of the urine as it flows through the nephron (i.e., changes in osmolality, pH and uric acid concentration). However, the presence of acanthocytes (i.e., RBCs in a donut form with one or more protruding cytoplasmic blebs) in urine is particularly noteworthy (Fig. 7.15). The conversion of RBCs into acanthocytes is not induced by changes in osmolality or pH. Rather, the physical forces undergone by RBCs as they pass through the glomerular filtration barrier (i.e., basement membrane) disrupt and permanently alter their cell membranes. Dysmorphic RBCs tend to be smaller, and often RBC fragments and other dysmorphic forms are present with acanthocytes. Studies indicate that when 5% or more of the RBCs in urine sediment are acanthocytes, it is an indicator of hematuria due to a glomerular disorder.1012 Rarely observed are sickle cells, which have been seen in the urine sediment of patients with sickle cell disease. Using phase contrast or interference contrast microscopy enhances the ability to evaluate RBC morphology and is recommended.

Table 7.5

Forms of Red Blood Cells in Urine

Form Name
Category Common Name Bessis Nomenclaturea Phase Microscopy Example Form Description
Isomorphic Normal cell Discocyte image Biconcave disk form of normal size
  Burr cell Echinocyte or crenated cell image Cell with evenly spaced projections or spicules over cell surface; this “reversible” shape change progresses from a “crenated” disk to a “crenated” sphere.10
  Ghost cell Ghost cell image Cell with thin membrane and without hemoglobin
Dysmorphic Acanthocyte or G1 cell Acanthocyte image Cell in a ring form (donut shape) with one or more cytoplasmic blebs (i.e., vesicle-shaped protrusions or bulges)
  Target cell Codocyte image Bull’s-eye appearance; can be bell- or cup-shaped
  Schistocyte Schizocyte image Cell fragment often with two or three pointed ends; size and shape vary
  Stomatocyte Stomatocyte image Cell with central pallor that appears slit-like; this shape change is “reversible.”10


aBessis M: Red cell shapes. An illustrated classification and its rationale. Nouvelle Revue Française d’Hématologie 12:721–746, 1972.

Normally, RBCs are found in the urine of healthy individuals and do not exceed 0 to 3 per high-power field or 3 to 12 per microliter of urine sediment.13 Semiquantitation is made by observing 10 representative high-power fields and averaging the number of erythrocytes seen in each. Although RBCs are nonmotile, they are capable of passing through pores only 0.5 mm (500 nm) in diameter.14 In addition, during inflammation, RBCs can be transported out of capillaries by the same mechanism as inert, insoluble substances.12 All RBCs in urine originate from the vascular system. The integrity of the normal vascular barrier in the kidneys or the urinary tract can be damaged by injury or disease, causing leakage of RBCs into any part of the urinary tract. Increased numbers of RBCs along with red RBC casts indicate renal bleeding, either glomerular or tubular. These urines also have significant proteinuria. When an increased number of RBCs is present without casts or proteinuria, the bleed is occurring below the kidney or may be caused by contamination (e.g., menstrual, hemorrhoidal).

Correlation With Physical and Chemical Examinations

RBCs observed during microscopic examination should be correlated with physical and chemical examinations (Table 7.6). Macroscopically, the urine sediment may indicate the presence of RBCs when the sediment button is characteristically red in color. Sometimes specimens have a positive chemical test for blood, but the microscopic examination reveals no RBCs. This can be explained by the fact that RBCs readily lyse and disintegrate in hypotonic or alkaline urine; such lysis can also occur within the urinary tract before urine collection. As a result, urine specimens can be encountered that contain only hemoglobin from RBCs that are no longer intact or microscopically visible. However, it is important to note that other substances, such as myoglobin, microbial peroxidases, and strong oxidizing agents, can cause a positive blood chemical test (see Chapter 6). Note that these reactions are considered “false-positive” reactions because RBCs or blood is not present.

Table 7.6

Red Blood Cells: Microscopic Features and Correlations
Microscopic features

Oct 18, 2022 | Posted by in PATHOLOGY & LABORATORY MEDICINE | Comments Off on Routine Urinalysis–the Microscopic Exam of Urine Sediment
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