MLSC 3054 Clinical Fluid Analysis

Charity Accurso, Ph.D.

Chapter 7

0-8 per HPF, or ~10 WBCs per μL is

acceptable
Increased numbers – inflammation
Kidneys to lower urinary tract
20+ WBCs per μL

 Basic test principle

 Azurophilic granules–neutrophils, eosinophils, basophils, monocytes,
macrophages
 Not present in lymphocytes
 Leukocyte esterases catalyze the hydrolysis of esters which liberate a
pyrrole. The pyrrole reacts with a diazonium salt to produce a purple
product.
 Terms
 Trace, mod, large
 Trace, 1+, 2+, 3+
 Confirmatory Test
 Microscopic confirmation

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MLSC 3054 Clinical Fluid Analysis

 False Positives  False Negatives

 Strong oxidizing agents or  Elevated glucose (>3 g/dL)
formalin  High specific gravity –
 Pigmented materials that crenation of WBCs will prevent
mask the color reaction esterase release
 Vaginal secretions  Drugs
 Tetracycline, gentamicin,
cephalosporin
 High protein (> 500 mg/dl)
 Limitations
 Sensitivity: manufacturer’s
guidelines
 Specificity: does not detect
lymphocytes

 Rapid screening for UTI

 Bladder (cystitis), renal pelvis, tubules, or both
 Routes:
 External contamination
 From the bloodstream
 Failure to treat
 Inflammatory process continues
 Complications: renal tissue damage, impairment
of renal function, hypertension and septicemia

 Bacteria convert nitrate to nitrite

 Basic test principle – Greiss reaction
 Diazotization of nitrite with an aromatic amine to
produce a diazonium salt, followed by an azo-
coupling reaction with an aromatic chromogen

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MLSC 3054 Clinical Fluid Analysis

 Limitations  False Negatives

 Sensitivity: Refer to  Very high levels of bacteria –
manufacturer’s guidelines further convert nitrite to
 Specificity: specific for nitrogen
nitrite interferences  Non-nitrate producing bacteria
and yeasts
 False Positives
 Low dietary nitrate (vegetarian
 Pigmented material in the diet)
urine
 Elevated ascorbic acid
 Improper collection or
storage  Insufficient retention time in
bladder to produce nitrite - use
first morning specimen

 Confirmatory tests
 Microscopic examination
 Microbiological workup – culture
 Most are gram negative bacilli
 Reference range: Negative

 Normal urine - 1-15 mg/dL or 150 mg per 24 hours

 Proteinuria – early renal disease
 Albumin is major serum protein found in normal
urine
 Low due to most being retained in glomerulus
 Others: microglobulins, Tamm-Horsfall
(uromodulin), prostatic, seminal and vaginal
proteins, secretory Immunoglobulin A

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MLSC 3054 Clinical Fluid Analysis

 Does not always mean renal disease!

 Need follow-up
 Group proteinuria into 3 categories:
 Prerenal
 Renal
 Postrenal

 Conditions that affect the plasma prior to reaching

the kidney
 Not indicative of renal disease
 Often transient – due to inc levels of LMW plasma
proteins (hgb, myoglobin, acute phase reactants)
 Inc filtration exceeds normal reabsorptive
capacity leading to overflow
 Normally not discovered during routine UA as
strips are specific for albumin

 Bence Jones Protein

 Multiple myeloma – proliferation of Ig producing
plasma cells
 Serum contains inc monoclonal Ig light chains –
Bence Jones protein
 Appears in urine when exceeds reabsorption
capacity

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MLSC 3054 Clinical Fluid Analysis

 When suspected, use screening test based on

unique solubility characteristics of protein
 Bence Jones protein does not remain coagulated
when heated – unlike other proteins
 Coagulates between 40-60°C and dissolves at
100°C
 Not all patients with MM excrete Bence Jones

 May be due to glomerular or tubular damage

 Glomerular membrane damage
 Impairment of selective filtration
 Inc serum albumin and eventually RBCs and WBCs in urine
 Causes: conditions that present the glomerulus with abnormal
substances – amyloid material, toxic substances, immune complexes
in SLE and streptococcal glomerulonephritis
 Increased pressure from blood entering glomerulus
 Overrides filtration resulting in albumin in urine
 May be reversible – dehydration or strenuous exercise
 Pre-eclampsia
 Protein levels: slightly above normal to 4 g/day

 Tubular damage
 Inc albumin b/c it cannot be reabsorbed by tubules
 Other LMW proteins also present
 Causes: exposure to toxic substances and heavy metals, severe
viral inf, Faconi’s syndrome
 Protein levels: not markedly elevated (as in glomerular
damage)
 Several benign causes of renal proteinuria – exposure to
cold, strenuous exercise, high fever, dehydration

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MLSC 3054 Clinical Fluid Analysis

 Orthostatic (Postural) Proteinuria

 Functional proteinuria
 Increase in urine protein excretion when the individual is in an
upright position
 Disappears when horizontal
 Cause: believed to be increased pressure on renal vein when
vertical
 Detection: collect specimen immediately after waking and again
after being vertical for several hours
 If condition present:
 First morning specimen: negative
 Standing specimen: positive

 Microalbuminuria – small amt of albumin

 Detected in onset of renal complications due to diabetes
 Indicates the need for better glucose control and control of
hypertension
 Not detected on reagent strip
 Reported as: albumin excretion rate (AER) in μg/min or
mg/24 h
 Albumin:creatinine ratio
 Significant: AER is 20-200 μg/min; 30-300 mg albumin
excreted in 24h or albumin:creatinine ratio is >3.4
mg/mmol

 AER and 24h albumin – timed specimens

 Albumin:creatinine ratio – random specimen
 Microalbumin reagent strips – measures semi-
quant both albumin and creatinine
 Micral test – reagent strip with antibody-enzyme
conjugate to detect albumin
 Can measure 0-10 mg/dL

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MLSC 3054 Clinical Fluid Analysis

 Protein added as it passes through lower urinary

tract (ureters, bladder, urethra, prostate, vagina)
 Exudates from bacterial and fungal inf contain
proteins
 Proteins from blood due to injury or menstrual
contamination, prostatic fluid, large amts of sperm

 Basic test Principle  Results: trace, 1+, 2+,

 Protein error of indicators-
3+, 4+
color change occurs in the  Semi-quant results also
presence of protein  Limitations
 pH maintained  Sensitivity: 20-30 mg/dL
 Proteins will result in release  Specificity: Albumin
of H+ from indicator dye
 Yellow in absence of protein

Interferences
 False Positives False Negatives
 Buffered or highly alkaline pH –  Proteins other than albumin
overrides acid buffer system  Very dilute urine
 Rise in pH causing color change not  Very high salt concentration
reflective of protein conc
 Gross hematuria
 Some skin cleansers/disinfectants
(High pH) Dyes and pigments
 Extended contact w/ urine
removing buffer

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MLSC 3054 Clinical Fluid Analysis

Additional tests
 Sulfosalicylic acid (SSA) – precipitation test – table 7.7, figure
7.3
 Reacts equally with all forms of protein
 Confirm protein in presence of highly alkaline urine
 Other substances precipitated by acid will interfere
 TCA – trichloroacetic acid
 Similar to SSA
 May also indicate presence of drugs
 Microalbumin

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