ANALYSIS OF URINE & BODY FLUIDS (MT 6328) LECTURE 

Asst. Prof. Ron Christian G. Sison, MPH | 1​ Shifting st​

 
RENAL FUNCTION TESTS 
● eGFR
OVERVIEW OF RENAL FUNCTION TESTS ● Cystatin C
● Beta​2​-Microglobulin
● Renal Function Tests  
○ Tests for GFR From the book:
○ Urinalysis (separate discussion) ● Clearance tests - standard tests used to measure the
○ Tests for Renal Tubular Acidosis filtering capacity of the glomeruli
○ Tests of Kidney Concentrating Ability
○ Measures the rate at which the kidneys are
● Summary
able to remove or clear a filterable substance
RENAL FUNCTION TESTS from the blood
○ To ensure that glomerular filtration is being
● Detect renal damage measured accurately, the substance analyzed
● Monitor functional damage must be one that is neither reabsorbed nor
● Help determine etiology secreted by the tubules.
● From a clinical perspective it is important to have test which
● Other factors to consider in selecting a clearance tests
would have these characteristics. No such test exists.
include:
● An early test to detect renal damage, for instance a simple
strip test for haematuria is important in screening for heavy ○ Stability of the substance in urine during 24-hr
metal poisoning. collection period
● There is a clinical need to monitor a patient with renal ○ Plasma level consistency
disease and this is achieved by serial plasma ○ Substance’s availability to the body
measurements. ○ Availability of tests to analyze the substance
● We need to know when to start dialysis in renal failure and
 
laboratory tests assist the clinical decision making.
● There are about a million nephrons in each kidney and this GLOMERULAR FILTRATION RATE 
represents a considerable functional reserve. In renal
disease about half the nephrons have to lose their - Volume of blood filtered across glomerulus per unit time
functioning before the abnormality can be detected by - Best single measure of kidney function
conventional laboratory tests. - Normally ​100-130 mL/min ​(120mL/min average)
- Determined by:
LABORATORY TESTS FOR RENAL FUNCTION ● Net filtration pressure across glomerular
basement membrane
1. Glomerular Filtration Rate (GFR) ● Permeability and surface area of glomerular
2. Plasma Creatinine basement membrane
3. Plasma Urea - Patient’s remain asymptomatic until there has been a
4. Urine Volume significant decline in GFR
5. Urine Urea - Can be very accurately measured using “gold standard”
6. Minerals in Urine technique
7. Urine Protein - Is an ​ideal marker
8. Urine glucose ○ Produced normally by the body (exogenous vs
9. Hematuria endogenous)
10. Osmolality ○ Produced at a constant rate
○ Filtered across glomerular membrane
The measurement of urine protein is important in certain conditions, ○ Removed from the body only by the kidney
e.g.diabetes. The detection of substances such as red cells or filtered only, not reabsorbed or secreted
glucose could be an early indicator of renal damage.
CANDIDATE MARKERS FOR GFR
Inulin  Filtered only
TESTS FOR GLOMERULAR FILTRATION RATE
Not made by body
Must be injected
● Urea
Creatinine  An endogenous product of muscle metabolism
● Creatinine
Near-constant production
● Creatinine Clearance
Filtered, but a bit secreted

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Urea  An endogenous product of protein intake Blood in GI tract
Filtered and absorbed
Synthesis varies with diet Why does BUN increase?
  ● Decrease GFR, but ​also​: 
From the book: ● Increased renal reabsorption:
● A test that requires an infused substance is termed an ○ ​ECV depletion
exogenous procedure and is seldom the method of ● Increased hepatic urea synthesis
choice if a suitable test substance is already present in ○ ​High protein feeding
the body (​endogenous procedure​) ○ Corticosteroid treatment (Prednisone, etc.)
○ GI blood absorption
HISTORICAL NOTE
BLOOD UREA NITROGEN (BUN) 
Urea Clearance
 
● The earliest glomerular filtration tests measured urea - Imperfect marker of decrease GFR
because of its presence in all urine specimens and the - Marker of adequacy of protein intake
existence of routinely used methods of chemical - Marker for presence of uremic toxins in chronic renal failure
analysis. - BUN:Crea ratio reflects ECV volume status:
● Because approximately 40% of the filtered urea is ● 10:1 = normal
reabsorbed, normal values were adjusted to reflect the ● >20:1 if ECV contracted.
reabsorption, and patients were hydrated to produce a ○ Increase proximal tubule Na and urea
reabsorption
urine flow of 2 mL/min to ensure that no more than 40%
of the urea was reabsorbed. CREATININE

Inulin Clearance  ● Product of muscle metabolism

● Inulin, a polymer of fructose, is an extremely stable ● Some creatinine is of dietary origin
substance that is not reabsorbed or secreted by the ● Freely filtered, but also actively secreted into urine
tubules. It is not a normal body constituent, however, ● Secretion is affected by several drugs
 
and must be infused by IV at a constant rate throughout
From the book:
the testing period.
The use of creatinine has several disadvantages and careful
● Therefore, although inulin was the original reference consideration should be given to them. They are as follows:
method for clearance tests, current methods are 1. Some creatinine is secreted by the tubules, and
available that are endogenous and can provide accurate secretion increases as blood levels rise.
GFR results. 2. Chromogens present in human plasma react in the
chemical analysis. Their presence, however, may help
UREA counteract the falsely elevated rates caused by tubular
secretion.
● It is used historically as a marker of GFR
● Freely filtered but both re-absorbed and excreted into the 3. Medications, including gentamicin, cephalosporins, and
urine cimetidine (Tagamet), inhibit tubular secretion of
● Re-absorption into blood increased with volume depletion; creatinine, thus causing falsely low serum levels.
therefore GFR underestimated 4. Bacteria will break down urinary creatinine if specimens
● Diet, drugs, disease all significantly affect Urea production are kept at room temperature for extended periods.
● Product of protein catabolism 5. A diet heavy in meat consumed during collection of a
● Filtered
● Reabsorbed in proximal tubule 24-hour urine specimen will influence the results if the
● If sodium is avidly reabsorbed, so is urea plasma specimen is drawn before the collection period
● Serum urea concentration is measured as ​Blood Urea  because the increased intake of meat can raise the urine
Nitrogen (BUN) and plasma levels of creatinine during the 24-hour
  collection period.
INCREASE  DECREASE  6. Measurement of creatinine clearance is not a reliable
Volume depletion  Volume Expansion indicator in patients suffering from muscle-wasting
Dietary protein  Liver expansion diseases or persons involved in heavy exercise or
Corticosteroids  Severe malnutrition
athletes supplementing with creatine.
Tetracyclines

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7. Accurate results depend on the accurate completeness LIMITATIONS OF CREATININE CLEARANCE
of a 24-hour collection. ● Only valid at steady state
8. It must be corrected for body surface area, unless ○ [Cr]serum must be stable
● Trimethoprim, cimetidine​ lower tubular Creatinine
normal is assumed, and must always be corrected for
secretion and lower Creatinine clearance without changing
children. GFR
○ Becomes more inaccurate at low GFR
● Newer methods that do not require the collection of
timed (24-hour) urine specimens have been developed ANOTHER PROBLEM WITH CREATININE CLEARANCE
using just the (1) serum creatinine, (2) cystatin C, or (3) ● Must be done on a properly collected, ​timed-sample​ -
patient error
beta2-microglobulin values. The results of these tests
● How can we check accuracy of any timed urine collection?
are reported as ​estimated glomerular filtration rate 
(eGFR)​. CREATININE EXCRETION
  ● The amount of creatinine excreted per day is stable for a
SERUM CREATININE CONCENTRATION  given patient
  ● It is function of ​muscle mass​: generally higher in
- Normally 0.7-1.4 mg/dl, depending on muscle mass ○ men vs. women
- Inversely proportional to GFR ○ youth vs elderly
- Good way to follow changes in GFR ● Expressed per kg lean body mass as the ​creatinine index 
- But also elevated by increase of muscle mass, decrease of
QUICK FORMULAE FOR ESTIMATING GFR
tubular secretion 
● Include some combination of sex, weight, serum creatinine,
Increase  Decrease  race, and age
Male  Female ● Use only at steady state (stable SCr)
Meat in diet  Age ● Useful screens for decreased GFR, esp. in elderly and
Muscular body type  Malnutrition small people, where errors in drug dosing may be major
Cimetidine & some other Muscle wasting
medications CREATININE TEST SUMMARY
Amputation

CREATININE CLEARANCE 
 
- Measure serum and urine creatinine levels and urine
volume, and calculate serum volume cleared of creatinine
- Same issues as with serum creatinine, except muscle
mass
- Requirements for 24-hour urine collection adds variability
and inconvenience
FORMULA:
CYSTATIN C

● It is a 13 kD protein produced by all cells at a constant rate

- Therefore, it represents the ​volume of serum completely  ● Freely filtered
cleared of creatinine per unit time ● Reabsorbed and catabolized by the kidney and does not
- Since virtually all creatinine is cleared via glomerular appear in the urine
filtration, it closely approximates the GFR
From the book:
EXAMPLE: ● Measurement of serum cystatin C has been shown to
UCr = 72 mg/dl provide a good procedure for screening and monitoring
SCr = 2.0 mg/dl GFR.
V = 2 liters ● Cystatin C 
time = 24 hours ○ Small protein produced at a constant rate by
all nucleated cells

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○ Readily filtered by the glomerulus and From the book: 
reabsorbed and broken down by the renal CLINICAL SIGNIFICANCE
tubular cells ● When interpreting the results of a creatinine clearance
○ Advantage​: independent of muscle mass test, the GFR is determined not only by the number of
● No cystatin C is secreted by the tubules, and the serum functioning nephrons but also by the ​functional ​capacity
concentration can be directly related to the GFR. of these nephrons.
● Immunoassay procedures are available for measuring ○ Even though half of the available nephrons
cystatin C. may be nonfunctional, a change in the GFR
● Monitoring levels of cystatin C is recommended for:   will not occur if the remaining nephrons double
○ Pediatric patients their filtering capacity.
○ Persons with diabetes ■ Evidenced by persons who live
○ The elderly normal lives with only one kidney
○ Critically ill patients ● Although the GFR is a frequently requested laboratory
● Recent studies also have shown that measuring ​both  procedure, its value does not lie in the detection of early
serum  or  plasma  cystatin  C  and  creatinine  ​can renal disease. Instead, it is used to:
provide even more accurate information on a patient’s ○ Determine the ​extent of nephron damage in
GFR. known cases of renal disease
○ Monitor  the  effectiveness  of  treatment 
BETA-2-MICROGLOBULIN designed to prevent further nephron damage
○ Determine the feasibility of administering 
● Molecular weight 11,800, dissociates from human medications​, which can build up to dangerous
leukocyte antigens at a constant rate blood levels if the GFR is markedly reduced.
● Is rapidly removed from the plasma by glomerular filtration
● Sensitive methods using​ enzyme immunoassay​ are
available for the measurement of beta2-microglobulin. ESTIMATED GLOMERULAR FILTRATION RATE
● A rise in the plasma level of beta2-microglobulin has been
shown to be a more sensitive indicator of a decrease in ● Increasing requirements for dialysis and transplant (8 –
GFR than creatinine clearance. 10% per year)
● The test is not reliable in patients who have a history of ● Shortage of transplantable kidneys
immunologic disorders or malignancy ( clinical correlation?) ● Large number at risk
  ● eGFR calculation has been recommended by National
RADIONUCLEOTIDES Kidney Foundation whenever a serum creatinine is
performed in adults
● Exogenous procedures; more labor intensive and costly  
● Injecting radionucleotides such as ​125​I- iothalamate Stage  Description  GFR  Prevalence​3 
provides a method for determining glomerular filtration ML/min/1.173m​ 2 

through the plasma disappearance of the radioactive 1 Kidney Damage >90 478,500
material and enables visualization of the filtration in one or with Normal or
both kidneys ↑ GFR
● This procedure can be valuable to measure the viability of 2 Kidney Damage 60 - 89t 435,000
a transplanted kidney. with Mild ↓ GFR
3 Moderate ↓ GFR 30 - 59 623,500
4 Severe ↓ GFR 15 - 29 29,000
5 Kidney Failure <15 0r dialysis 14,500
 
Cumulative 8-year mortality rate, depending on serum
creatinine level at baseline, in the Hypertension Detection
and Follow-up Program 
Serum creatinine mg/dL Mortality rate (%)
(umol/L)
0.8 - 0.99 (71 - 88) 10
1.1 - 1.29 (97 - 114) 12
1.3 - 1.49 (115 - 132) 16
1.5 - 1.69 (133 - 149) 22

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1.7 - 1.99 (150 - 176) 30 ○ High molecular weight proteins are retained in
2.0 - 2.49 (177 - 220) 41 the circulation by the glomerular filter (Albumin,
≥ 2.5 (≥ 221) 54 Immunoglobulins)
○ Low molecular weight proteins are filtered then
● Problem: reabsorbed by renal tubular cells
○ Need an easy test to screen for early decreases ● ​Glomerular​:
in GFR that you can apply to a large, at-risk ○ Mostly albumin, because of its high concentration
population and therefore high filtered load
○ Can serum creatinine be made more sensitive by ● Tubular​:
adding more information? ○ Low molecular weight proteins not reabsorbed by
tubular cells (e.g. alpha-1 microglobulin)
● Overflow​:
eGFR by MDRD Formula
○ Excessive filtration of one protein exceeds
reabsorptive capacity (BenceJones, myoglobin)
● Mathematically modified serum creatinine with additional
information from patients age, sex and ethnicity ALBUMIN CREATININE RATIO (MICROALBUMIN)
● eGFR = 30849.2 x (serum creatinine)-1.154 x (age)-0.203
(if female x (0.742)) ● Normal albumin molecule
● In health, there is very little or no albumin in the urine
The MDRD-IDMS traceable formula is: ● Most dip sticks report albumin at greater than 150 mg/L

GFR = 175 x serum creatinine​-1154​ x age​-0.203​ x 0.742 URINARY ALBUMIN

(if patient is female) x 1.202 (if patient is black)
● Detection of low levels of albumin (even if below dipstick
*Creatinine filtration and Secretion: cut-off) is predictive of future kidney disease with diabetes
● Very significant biologic variation usually requires repeat
collections
● Treatment usually based on timed urine albumin collections

FOLLOW-UP BASED ON SCREEN RESULTS

● Kidney Ultrasound
● Specialist Referral
● Cardiovascular Risk Assessment
● Diabetes Control
● Smoking cessation
● Hepatitis / Influenza Management

CREATININE STANDARDIZATION

● Based on Isotope dilution /mass spectrometry

measurements of creatinine standards
SCREEN HIGH RISK GROUPS ● Permits estimation and correction of creatinine and eGFR
bias at the laboratory level
1. eGFR
2. Urinalysis IMPORTANCE OF STANDARDIZATION
3. Albumin / Creatinine Ratio
Low bias  ● Causes inappropriately increased eGFR
TESTS THAT PREDICT KIDNEY DISEASE creatinine  ● Patients will not receive the benefits of
more intensive investigation of treatment
1. eGFR High bias  ● Causes inappropriately decreased eGFR
2. Albumin Creatinine Ratio (aka ACR or Microalbumin) creatinine  ● Patients receive investigations and
treatment which is not required
PROTEINURIA ● Wastes time, resources and increases
anxiety
● In health:  
POOR CREATININE PRECISION

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From the book: 
● Incorrect categorization of patients with both “normal” and ● Concentration tests 
decreased eGFR. ○ Tests to determine the ability of the tubules to
reabsorb the essential salts and water that
TOTAL ERROR
have been nonselectively filtered by the
● TE = % bias + 1.96 CV glomerulus
● Goal is <10% (requires bias ≤ 4% and CV ≤ 3%) ● Throughout the years, various methods have been used
to produce water deprivation, including the Fishberg 
KIDNEY FUNCTIONS and Mosenthal concentration tests​, which measured
specific gravity.
● Selectively secretes into or re-absorbs from the filtrate to ● Fishberg Test 
maintain
○ Patients were deprived of fluids for 24 hours
 
before measuring specific gravity
  Tests 
Water Balance  • specific gravity ● Mosenthal Test 
• osmolarity ○ Compared the volume and specific gravity of
• water deprivation testing day and night urine samples to evaluate
• Antidiuretic hormone concentrating ability
Retention of nutrients   • Proteins ● Neither test is used now because the information
• Sugar provided by ​specific gravity measurements is most
• Amino acids
useful as a screening procedure, and quantitative
• Phosphate
measurement of renal concentrating ability is best
Secretes waste products  • Urate
• Oxalate assessed through ​osmometry​.
• Bile salts ● Currently renal concentrating testing is performed after
Salt Balance  • Na​+​, Cl​- various periods of fluid deprivation, measuring urine and
• K​+​ Aldosterone often serum osmolality.
• Renin ● Controlled intake procedures can include after dinner
Acid Base Balance  • pH overnight deprivation of fluid for ​12 hours followed by
• HCO​3-
collection of a urine sample.
• NH​4+​ Acid loading
• Urinary Anion Gap ○ Normal urine test​: a urine osmolality reading
of 800 mOsm or higher; test can be
TO SUMMARIZE discontinued
■ A urine to serum ratio (U:S ratio) of
1. Use the ​Creatinine Clearance​ as the best estimate of 3:1 or greater or a urine osmolality
GFR of 800 mOsm or greater indicates
2. Use the ​Serum Creatinine​ to follow renal function over normal tubular reabsorption.
time ○ Abnormal urine test: ​the fluid is restricted for
3. Use the ​Creatinine Index​ to check the adequacy of a urine another two hours and both urine and serum
collection species are collected for osmolality testing.
4. Use the​ BUN​ to help assess GFR, volume status, and ■ If the test continues to be abnormal,
protein intake additional testing is performed to
determine whether the failure to
TESTS OF KIDNEY CONCENTRATING ABILITY concentrate the urine is caused by
diabetes insipidus that occurs as the
● To differentiate
result of a problem with the
● Psychogenic polydipsia
production or the response of the
● Central diabetes insipidus
● Nephrogenic diabetes insipidus kidney to ADH.
■ The patient is ​injected  ​with ADH
TUBULAR REABSORPTION TESTS and serum and urine specimens are
collected in 2 and 4 hours. If at this
time the test is ​normal​, it indicates

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that the patient is not capable of ● Therefore osmolality is performed for a more accurate
producing ADH (neurogenic evaluation of renal concentrating ability.
diabetes insipidus) and if the test is
abnormal ​then the renal tubules are FREEZING POINT DEPRESSION OSMOMETER
not responding to ADH (nephrogenic
diabetes insipidus).  ● Principle: ​the freezing point of a solution is related to the
osmotic concentration of that solution.
● If concentration of the solute is increased, it will lower its
DETERMINATION OF PLASMA AND URINE OSMOLALITY freezing point.
● Normal Pure solvent (water) freezing point is ​- 0.53​°​C  
● Major osmotic substances in normal plasma are ​sodium,  ● Most commonly used method for measuring the osmolality
chloride, glucose and urea
of serum or urine in clinical laboratory
● Osmolal gap​ - is difference between the measured
osmolality and calculated osmolality
● Theoretically all ​four colligative properties​ could be used
as basis to measure osmolality

TERMINOLOGIES

1. Osmotic Pressure ​- is the minimum pressure which needs to

be applied to a solution to prevent the inward flow of water
across a semipermeable membrane
- Also defined as the measure of the tendency of a solution
to take in water by osmosis  
2. Osmolarity​ - is defined as the number of osmoles of solute per From the book:
liter (L) of solution (osmol/L or Osm/L) ● These osmometers determine the freezing point of a
3. Osmolality ​- is a measure of the osmoles (Osm) of solute per solution by ​supercooling  a  measured  amount  of 
kilogram of solvent (osmol/kg or Osm/kg) sample  to  approximately  27°C​. The supercooled
- It is thermodynamically more exact expression because sample is vibrated to produce crystallization of water in
solution concentration expresses on weight basis are
the solution.
temperature independent;​ whereas those based on
volume vary with temperature ● The ​heat of fusion produced by the crystallizing water
- Is what the clinical laboratory measures temporarily raises the temperature of the solution to its
freezing point.
APPLICATION ● A temperature-sensitive probe called a ​thermistor​, in
which resistance decreases as temperature increases,
NORMAL VALUES: measures this temperature increase, which corresponds
● Urine osmolality: to the freezing point of the solution, and the information
○ 24-hr specimen = 300-900 mOsm/kg of H2O is converted into milliosmoles.
○ Random sample = 50-1200 mOsm/kg of H2O
● Osmolarity can be calculated by comparing the freezing
○ After 12hr fluid restriction = >850 mOsm/kg of
H2O point depression of an unknown solution with that of a
● Serum Osmolality:  known molal solution.
○ Adult = 280-303 mOsm/kg of H2O ● Clinical osmometers use solutions of known ​NaCl 
○ Newborn = upto 266 mOsm/kg of H2O concentration as their reference standards because a
● Urine-Serum Ratio: 1:1 to 3:1  solution of partially ionized substances is more
  representative of urine and plasma composition​.
From the book:
● Osmolality - measures only the number of particles in a VAPOR PRESSURE OSMOMETER
solution, whereas specific gravity is influenced by the
number and density (molecular weight) of the particles. ● Measurement related to the decrease in dew point of
● Renal concentration is concerned with small particles, temperature of pure solvent (water) caused by decrease in
primarily sodium and chloride molecules. vapor pressure of solvent by the solute.
● Large molecular-weight molecules such as glucose and ● Drawback​: measurement of any volatile solute in serum not
better.
urea do not contribute to the evaluation of renal
concentration.

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○ Volatile gases if present will increase the vapor From the book:
pressure of solvent) Factors to consider because of their influence on true osmolarity
● Hence, NOT recommended for clinical laboratory readings include:
● Lipemic serum 
○ The serum water displacement by insoluble
lipids produces erroneous results with both
vapor pressure and freezing point
osmometers.
● Lactic acid 
○ Falsely elevated values owing to lactic acid
formation also occur with both methods if
serum samples are not separated or
 
refrigerated within 20 minutes.
From the book:
● Vapor Pressure Osmometers  ● Volatile substances, such as ethanol
○ Dewpoint (temperature at which water vapor ○ Vapor pressure osmometers do not detect the
condenses to a liquid) presence of volatile substances, such as
alcohol, as they become part of the solvent
○ The depression of dew point temperature by
solute parallels the decrease in vapor phase
pressure, thereby providing its measurement. ○ However, measurements performed on similar
specimens using freezing point osmometers
○ are used primarily to analyze serum and sweat
microsamples for disorders not related to renal will be elevated.
function, such as cystic fibrosis. They are used
primarily in the chemistry department.
CLINICAL SIGNIFICANCE
 
● Procedure
From the book:
○ Samples are absorbed into small filter paper CLINICAL USES OF OSMOLARITY
disks that are placed in a sealed chamber
● Initially evaluating renal concentrating ability
containing a temperature-sensitive
● Monitoring the course of renal disease
thermocoupler. The sample evaporates in the
● Monitoring fluid and electrolyte therapy
chamber, forming a vapor.
● Establishing the differential diagnosis of ​hypernatremia
○ When the temperature in the chamber is and ​hyponatremia
lowered, water condenses in the chamber and
● Evaluating the secretion of and renal response to ADH
on the thermocoupler.
○ The heat of condensation produced raises the REFERENCE OSMOLALITY VALUES
temperature of the thermocoupler to the dew
● SERUM: Range from ​275-300 mOsm 
point temperature. This dew point temperature
● URINE: Range from ​50-1400 mOsm 
is proportional to the vapor pressure from the
○ Factors such as ​fluid intake and ​exercise can
evaporating sample. Temperatures are
greatly influence the urine concentration
compared with those of the NaCl standards
● Under normal random conditions, the ratio of urine to
and converted into milliosmoles.
serum osmolality should be at least ​1:1
○ The vapor pressure osmometer uses
○ After controlled fluid intake: should reach ​3:1 
microsamples of less than 0.01 mL; therefore,
● The ratio of urine to serum osmolality, in conjunction
care must be taken to prevent any evaporation
with procedures such as controlled fluid intake and
of the sample prior to testing.  
injection of ADH, is used to differentiate whether
diabetes insipidus is caused by decreased ADH
TECHNICAL FACTORS
production or inability of the renal tubules to respond to
  ADH.
○ Failure to achieve a ratio of 3:1 after injecting
ADH indicates that the collecting duct does not
have functional ADH receptors. In contrast, if
concentration takes place after ADH injection,

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an inability to produce adequate ADH is From the book:
indicated.  ● Tests to measure tubular secretion of nonfiltered
substances and renal blood flow are closely related in
OVERNIGHT WATER DEPRIVATION TESTING that total renal blood flow through the nephron must be
measured by a substance that is ​secreted ​rather than
● (Serum osmolarity <295 monitor patient weight hourly) filtered through the glomerulus.
● Collect urine hourly from 06:00 for osmolarity ● Impaired  tubular  secretory  ability or ​inadequate 
● Baseline serum osmolarity, Na+, ADH presentation  of  the  substance  to  the  capillaries
● When osmolarity plateaus repeat above tests and owing to decreased renal blood flow may cause an
administer ADH abnormal result.
● The test most commonly associated with tubular
INTERPRETATION secretion and renal blood flow is the p-aminohippuric 
acid (PAH) test. 
● If urine concentrates (osmolarity >600 and serum
osmolarity below <295)
● Normal physiology (Psychogenic polydipsia) PARA-AMINOHIPPURATE (PAH)

● Undergoes tubular secretion and is freely filterable.

○ At ​low [PAH]p​l, virtually all PAH escaping
filtration is secreted by the tubule.
● Therefore virtually all ​plasma supplying secreting 
nephrons ​is cleared of PAH.
● About​ 10-15% of total renal plasma flow (TRPF) ​supplies
non-secreting portions of the kidney. Thus, ​CPAH​
​ ​actually
measures ​effective renal plasma flow (ERPF)​.
● 8% of the renal blood flow does not come into contact with
the functional renal tissue
● This makes ​~ 85-90% of TRPF​.
 
From the book:
● To measure the exact amount of blood flowing through
the kidney, it is necessary to use a substance that is
completely removed from the blood (plasma) each time
● No urine concentration
○ Positive response to ADH it comes in contact with functional renal tissue.
■ Central diabetes insipidus ○ The principle is the same as in the clearance
○ No response to ADH test for glomerular filtration
■ Nephrogenic insipidus ○ However, to ensure measurement of the blood
flow through the entire nephron, the substance
must be removed from the blood primarily in
the peritubular capillaries rather than being
removed when the blood reaches the
glomerulus.
● Although it has the disadvantage of being exogenous,
the ​chemical  PAH  ​meets the criteria needed to
measure renal blood flow.
○ This nontoxic substance is loosely bound to
plasma proteins, which permits its complete
TUBULAR SECRETION AND RENAL BLOOD FLOW TESTS 
removal as the blood passes through the
 
peritubular capillaries
○ Except for a small amount of PAH contained in
plasma that does not come in contact with
functional renal tissue, all the plasma PAH is
secreted by the proximal convoluted tubule.

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○ Therefore, ​the  volume  of  plasma  flowing  = U​PAH​ V 
through  the  kidneys  determines  the   
amount of PAH excreted in the urine. 
● Standard clearance formula ​used to calculate effective
renal plasma flow:

● Based on normal hematocrit readings, reference values

for the effective renal plasma flow range from ​600 to 700 
mL/min,  ​making the average renal blood flow about
1200 mL/min. 
● The actual measurement is renal plasma flow rather
than renal blood flow, because the PAH is contained
only in the plasma portion of the blood. 2. The rate at which PAH enters the kidney is equal to the
rate that it leaves. Then,
○ Also, the term “effective” is included because
approximately 8% of the renal blood flow ​does 
not come into contact with the functional renal
tissue.
● The amount of PAH infused by IV must be monitored
3. Note that this last expression is the ​Direct Fick formula​ for
carefully to ensure accurate results; therefore, the test is
measuring plasma flow: the blood flow is equal to the rate
usually performed by specialized renal laboratories. of consumption (excretion) divided by the arterio-venous
● Nuclear medicine procedures using ​radioactive  concentration difference.
hippurate can determine renal blood flow by measuring 4. It is common to make the approximation that the
the plasma disappearance of a single  ​radioactive concentration of PAH in renal venous blood is zero.
injection and at the same time provide visualization of Substituting zero for venous [PAH], allows us to compute a
quantity called the effective renal plasma flow (ERPF).
the blood flowing through the kidneys. 

ESTIMATION OF RENAL PLASMA FLOW RATE:

● Certain substances are extracted from peritubular capillary 5. Note that the ERPF is equal to the clearance of PAH and
plasma and secreted into the proximal tubular fluid in large that it underestimates the true renal plasma flow by
amounts. approximately 10%.
● The concentration of such substances in renal venous 6. The "normal value" of ERPF in a 70 kg human is about 625
plasma is therefore, much less than in renal arterial ml/min. Since the PAH clearance underestimates renal
plasma. plasma flow by about 10%, the true renal plasma flow
● In such cases, the direct Fick Principle (blood flow = rate of (RPF) is about 700 ml/min.
excretion/(A-V) concentration difference) can be used to 7. The renal blood flow (RBF) is then RBF = RPF/1 - Hct ml
estimate renal plasma flow rate. blood/min, so that if the hematocrit is 45%, and RPF = 700
● PAH (para-aminohippuric acid) might be used for this ml/min, then RBF is 1273 ml/min. This is 20 to 25% of
purpose. cardiac output.

1. In the steady state, the rate at which PAH enters the From the book:
kidney (moles/min) in the renal arterial plasma is equal to HISTORICAL NOTE:
the rate at which PAH leaves the kidney in the urine and in ● Phenolsulfonphthalein Test 
renal venous blood. ○ Historically, excretion of the dye
○ PAH (moles/min) entering kidney in renal  phenolsulfonphthalein (PSP) was used to
arterial plasma = P​a​PAH ​x RPF  evaluate these functions. Standardization and
interpretation of PSP results are difficult,
○ PAH (moles/min) leaving kidney in renal  however, because of interference by
venous plasma = P​ ​v​ ​PAH​ ​x RPF  medications, elevated waste products in
.  patient’s serum, the necessity to obtain
○ PAH (moles/min) leaving kidney in the urine   several very accurately timed urine

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 MT 6328: ANALYSIS OF URINE AND BODY FLUIDS | LECTURE | 1​st​ SHIFTING 
 
specimens, and the possibility of producing
anaphylactic shock. Therefore, the PSP test is
not currently performed.   

TITRATABLE ACIDITY

● A normal person excretes approximately​ 70 mEq/day of 

acid ​in the form of titratable acid (H+), hydrogen phosphate
ions (​H2PO4–​), or ammonium ions (NH4+).
● In normal persons, a diurnal variation in urine acidity
consisting of alkaline tides appears shortly ​after arising 
and ​postprandially ​at approximately ​2 p.m. and 8 p.m. 
● ​The lowest pH is found at night. (​Nocturnal acidosis​)
 
From the book:
● The ability of the kidney to produce acid urine depends
on the tubular secretion of hydrogen ions and production
and secretion of ammonia by the cells of the distal
convoluted tubule. 

RENAL TUBULAR ACIDOSIS

 
● The inability to produce an acid urine in the presence of
metabolic acidosis
● This condition may result from impaired tubular secretion of
hydrogen ​ions associated with the ​proximal convoluted 
tubule​ or defects in ​ammonia secretion​ associated with
the ​distal convoluted tubule​.

Tests for Renal Tubular Acidosis 

● Urinary Anion Gap
○ (Na+ + K+) – Cl-
● In acidosis the kidney should excrete NH4+ and the gap
will be negative
● If NH4 + is not present (or if HCO3 - is present) the gap will
be neutral or positive, implying impaired kidney handling of
acid load.
○ Urine Anion Gap = (Na+ + K+) –Cl-
● Ammonium Chloride Loading
○ Load with ammonium chloride
○ Hourly measurements of urine pH
○ Normal at least one pH below 5.5

URINE TITRATABLE ACIDITY

● Urine pH, titratable acidity, and urinary ammonia

measurements can be used to determine the defective
function.
● The tests can be run simultaneously on either fresh or
toluene- preserved urine specimens collected at 2-hour 
intervals from patients who have been primed with an acid
load consisting of ​oral ammonium chloride.
● By titrating the amount of free H+ (titratable acidity) and
then the total acidity of the specimen, the ammonium
concentration can be calculated as the difference between
the titratable acidity and the total acidity.

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