Routine Chemical Examination of Urine Flashcards
Routine Chemical Examination of Urine includes what observation of 3 components?
- pH
- Protein
- Glucose
Plastic strip that contains one or more chemical-impregnated absorbent pads
AKA dipstick
Reagent Strip
Familiarize the Reagent Strip Testing Procedure
- Mix the specimen
- Insert reagent strip
- Remove excess urine
- Time according to manufacturer’s directions
- Compare reaction colors with the manufacturer’s chart under a good light source
- Perform backup tests when indicated
- Be alert for the presence of interfering substances
- Understand the principles and significance of the rest; read package inserts
- Relate chemical findings to each other and to the physical and microscopic urinalysis results
Reagent Strip Testing Procedure
refrigerated specimens to reach what temp prior to testing ?
Room temperature
Reagent Strip Testing Procedure
For how many seconds should the reagent strip be dipped?
1 sec
Reagent Strip Testing Procedure
How should the reagent strip be dipped?
complete but briefly
Reagent Strip Testing Procedure
How to remove excess urine in reagent strip?
- Run edge of strip along rim of tube
- Blot edge on absorbent paper
Reagent Strip Testing Procedure
For how many seconds should you wait to read glucose after dipping reagent strip into urine?
30 seconds
Reagent Strip Testing Procedure
From what direction do you read the reagent strip?
Bottom to top
Reagent Strip Testing Procedure
Do you read glucose or leukocytes first? why?
Glucose becasue it reacts rapidly
T or F
Reading leukocytes first causes false positive results for glucose
F (false negative results)
Reagent Strip Testing Procedure
After waiting for the reagents to react with the strip, what should you do?
Compare reaction colors with the manufacturer’s chart under a good light source
Reagent Strip Testing Procedure
In comparing reaction color’s with manufacturer’s chart, in whwat direction should you hold the strip?
Horizontally close to the color chart (not touching)
T or F
Holding the reagent strip close to the manufacturer’s bottle to the point of contact should not be done since it results to contamination
T
Reagent Strip Testing Procedure
Where you will insert the strip then operate on for the machine to read it to you
Reagent strip reader
T or F
the MT should always perform backup test when indicated
T
Reagent Strip Testing Procedure
Backup test for glucose
Benedict’s test
Reagent Strip Testing Procedure
Backup test for protein
heat and acetic acid or 3% Sulfosalicylic acid TEST (Exton’s)
T or F
One should be alert for the presence of interfering substances
T duh
Reagent Strip Testing Procedure
You understand the principle and significance of the test by?
reading package inserts
Reagent Strip Testing Procedure
Relate ………. to each other and to the ………. ………….and ………… urinalysis results
pretend the period are blanks, di ko mapalce maayos yung blanks
Relate chemical findings to each other and to the** physical** and microscopic urinalysis results
T or F
Relating physical and miscroscopic results means that once color of urine is red, microscopically there should be RBC present
T
Care of Reagent Strips
How should reagent strips be stored?
- Store with desiccant in opaque tightly capped containers
- Store below 30 c
- Keep strips in original container
- Remove immediately prior to use
Care of Reagent Strips
Presence of air in reagent strips cause?
discoloration
Care of Reagent Strips
Where should the reagent strips be stored? at what temperature?
- with desiccant in opaque tightly capped containers
- below 30 c
Care of Reagent Strips
Do not expose the strips to?
volatile fumes
Care of Reagent Strips
Do not touch the ?
test areas
Care of Reagent Strips
Do not use if the chemical pads become?
discolored
T or F
One should remain to use if chemical pads become discolored
F (one should discard)
Care of Reagent Strips
The reagent in the strip that is usually the first one to be discolored
Urobilinogen
T or F
Do not use past the expiration date
T.
Care of Reagent Strips
Do not transfer test strips to ?
another vial
T or F
Touch the test zones with your fingers
F (DO NOT touch test zones with your fingers; hold bottom without test)
What are the 4 sources of error
- Unmixed specimen
- Strip in urine for extended period
- Excess urine in the strip
- Refrigerated specimen
Sources of error
These 2 tests are usually affected by unmixed specimen
RBCs and WBCs
T or F
Unmixed specimen cause RBCs and WBCs to precipitate at bottom of container which causes false (+) results
F (cause false negative result)
Sources of error
reagents will leech (lalabas), goes into the urine → no reagent in test pad
Strip in urine for extended period
Sources of Error
What is the required component in a reagent strip which is a test fro protein that acts as a buffer
Citric acid
Sources of Error
No citric acid causes what result?
False positive result
Sources of Error
Run-over between chemicals producing distortion of colors
Excess urine in the strip
Sources of Error
What should be done to avoid excess urine in the strip ?
- Blot edge of the strip
Sources of Error
Refrigerated specimen cause what results?
False negative results
since enzymes do not react at cold temp.
Do enzymes thrive in cold or hot environment?
hot environment; do not react in cold temperature
T or F
Glucose gives false negative results once strip is subjected to cold temperature
T
Familiarize Quality Control for Routine Chemistry
- Test open bottles of reagent strips with known positive and negative controls every 24 hours
- Resolve control results that are out of range by further testing
- Test reagents used in backup tests with positive and negative controls
- Perform positive and negative controls on new reagents and newly opened bottles of reagent strips
- Record all control results and reagent lot numbers
Quality control
Test open bottles of reagent strips using what method?
positive and negative controls
Quality Control
How often should one test open bottles of reagent strips with known positive and negative controls?
every 24 hours (once daily)
Quality control
What should one do when control results are out of range ?
Resolve by further testing or repeating it
Quality control
What are the following that should be tested for positive and negative control?
- Open bottle of reagent strip
- Test reagents in backup test
- New and newly opened bottles of reagent strips
Quality Control; T or F
one should record only the control results of the newly opened bottles
F (all control results and their reagent lot numbers)
Quality Control
What 2 things should be recorded?
All control results and reagent lot numbers
If you see this card, study the procedure for quality control of reagent strips
go
Test using different reagents or methodologies to detect the same substances as detected by the reagent strips
Confirmatory Test
Confirmatory Test
To what products are confirmatory tests done?
Tablets and liquid chemicals
Confirmatory Test
reflects the ability of the kidney to maintain normal hydrogen ion concentration in plasma and extracellular fluid
pH
pH; T or F
Secretion of H+ ions in urine hekp maintain acid-base balance in the body
T
pH
must buffer and eliminate excess acids
Blood
Buffering capacity of blood depends on?
bicarbonate ions (HCO3)
Secretion of hydrogen ions causes reabsorption of?
bicarbonates
acidosis, normal, alkalosis?
pH of 7.40
Normal
acidosis, normal, alkalosis
pH of 7.00
acidosis
acidosis, normal, alkalosis
8.75
alkalosis
pH range of a first morning specimen
5.0 – 6.0
pH range of a random specimen
4.5 – 8.0
pH of an after meal specimen
Alkaline
Familiarize the factors which could affect urine pH
- Acid base content of the blood
- Patient’s renal function
- UTI
- Patient’s dietary intake
- Age of the specimen
factors which could affect urine pH
What is the effect of UTI in urine?
increases alkalinity
factors which could affect urine pH
What is the effect of old age of specimen?
specimen becomes alkaline
Familiarize the Clinical Significance of urine pH
- Respiratory or metabolic acidosis
- Respiratory or metabolic alkalosis
- Defects in renal tubular secretion and reabsorption of acids and bases (renal tubular acidosis)
- Precipitation of crystals and calculi formation
- Treatment of UTI
- Determination of unsatisfactory specimen
Clinical Significance of urine pH
happens when lungs are unable remove enough CO2, leads to too much CO2 in body
metabolic acidosis
Clinical Significance of urine pH
Too much CO2 causes pH to become?
acidic
Clinical Significance of urine pH
Low CO2
metabolic alkalosis
Clinical Significance of urine pH
Tubules unable to reabsorb bicarbonates
Defects in renal tubular secretion and reabsorption of acids and bases (renal tubular acidosis)
Clinical Significance of urine pH
What happens if tubules cannot reabsorb bicarbonates?
goes into urine and will not secrete H+
Clinical Significance of urine pH
Precipitation of crystals and calculi formation depends on?
pH
Clinical Significance of urine pH
Crystals come from precipitation of what?
precipitation of inorganic chemicals
Clinical Significance of urine pH
What happens if crystals are not flushed out
it turns into stones
Clinical Significance of urine pH
most frequent constituent of renal calculi
Calcium oxalate
Clinical Significance of urine pH
In what pH of urine does the calcium oxalate precipitate?
acidic urine
Clinical Significance of urine pH
Since calcium oxalate precipitate in acidic urine, what do the physicians do in order to idscourage formation of calculi?
they alkalinize it
Clinical Significance of urine pH
pH which indicates specimen is already stale?
9.0
Clinical Significance of urine pH; T or F
Specimen should be recollected if it has pH of 9.0
T (stale already)
Acid or Alkaline Urine?
Increased protein and meat diet
Acid Urine
Acid or Alkaline Urine?
Cranberries
Acid Urine
Acid or Alkaline Urine?
Acid producing bacteria
Acid Urine
Acid or Alkaline Urine?
Starvation
Acid Urine
Acid or Alkaline Urine?
Dehydration
Acid Urine
Acid or Alkaline Urine?
Diarrhea
Acid Urine
Acid or Alkaline Urine?
Diabetes mellitus
Acid Urine
Acid or Alkaline Urine?
Increased fruits and vegetables
Alkaline Urine
Acid or Alkaline Urine?
Citrus
Alkaline Urine
Acid or Alkaline Urine?
after meal
Alkaline Urine
Acid or Alkaline Urine?
Renal tubular acidosis
Alkaline Urine
Acid or Alkaline Urine?
Urease producing bacteria
Alkaline Urine
Acid or Alkaline Urine?
Hyperventilation
Alkaline Urine
rabid release of carbon dioxide
Acid or Alkaline Urine?
Old specimen
Alkaline Urine
used as home remedy for urine
Cranberries
the common of these 4 are the formation of ketone bodies (presence of acid substances)
- Starvation
- Dehydration
- Diarrhea
- Diabetes mellitus
T or F
Hyperventilation causes rabid reabsorption of carbon dioxide hence produces acid urine
CONGRATS YOU’RE ON THE 100TH CARD
CELEBRATORY TWERK PLS SEND TO ME VID
F (opposite, hyperventilation is rabid release of CO2 hence alkaline urine)
What are the methods used to measure pH?
- pH strip
- pH electrode
- Titratable acidity
Methods of Measuring pH
- Double indicator system
pH reagent strip
Methods of Measuring pH
Indicators used in pH reagent strip
Methyl red & Bromothymol blue
Methods of Measuring pH
Principle of pH Reagent strip
Double indicator system
Methods of Measuring pH
Methyl red changes color from? also what pH range
red to yellow
pH 4-6
Methods of Measuring pH
Bromothymol blue changes from what color? also what pH
yellow to blue
pH 6-9
Methods of Measuring pH
In reagent strips, what pH does correspond to color orange?
5.0
Methods of Measuring pH
In reagent strips, what pH does correspond to color blue
8.5
Methods of Measuring pH
What is the color transition in reagent strip?
5.0 orange → green → blue (8.5) as pH ↑
Methods of Measuring pH
Reagent strips should be read after?
after 60 seconds
Methods of Measuring pH
- pH meter w/ glass electrode
- Measures the voltage caused by the H+ ions in the urine
- An electrode is dipped into the solutions
pH electrode
Methods of Measuring pH
Measured by titrating an aliquot of 24 hour urine with 0.1 N NaOH with pH 7.4 as an end point
Titratable Acidity
Methods of Measuring pH
What urine specimen is used in titratable acidity?
24 hour urine specimen
Methods of Measuring pH
In titratable acidity, what other substance are used with 24 hour urine?
0.1 N NaOH
Methods of Measuring pH
In titratable acidity, what is the pH end point?
pH 7.4
Important renal marker
Protein
Volume of clinical proteinuria?
> 30 mg/dl
T or F
Most of the albumin is not filtered
T
Filtered albumin is reabsorbed or excreted?
reabsorbed by tubule
Normal amount of protein in the body per day?
less than 100 mg of protein / 24 hours (150 from henry)
or
less than
10 mg/dl average random
According to henry’s and bishop, what is the molecular weight of protein allowed by glomerulus to filter?
Henry: < 70,000 mw
Bishop: < 66,000 mw
What are the other proteins in urine?
- Microglobulin
- Uromoduli
Other proteins in urine
What is the other name of uromodulin?
Tamm-Horsfall glycoprotein
Other proteins in urine
- Tamm-Horsfall glycoprotein
- lines matrix of all casts
Uromodulin
How many g/day is the protein content for functional proteinuria?
0.5 g/day
- 0.5 g/day
- This is usually resolved with rest for 2-3 days
Functional Proteinuria
How many days should one rest to resolved functional proteinuria
2-3 days
repeat collection after
Familiarize the causes of functional proteinuria
- Dehydration
- Exercise
- Congestive Heart Failure
- Cold exposure (prolonged cold baths)
- Fever
- Late pregnancy
- Emotional stress
- Exaggerated Lordotic posture * (Exaggerated lumbar curve)*
- Happens when patient is standing
Postural/Orthostatic
Positive or negative in urine protein?
Day
positive
Positive or negative in urine protein?
Night
negative
Positive or negative in urine protein?
Vertical positioning of the body
positive
Positive or negative in urine protein?
Horizontal positioning of the body
negative
Positive or negative in urine protein?
First voided urine
negative
Positive or negative in urine protein?
2 hrs standing or walking
positive
Familiarize the procedure in collecting sample postural/orthostatic
- Let patient urinate before bedtime and get sample
- Patient sleeps horizontally
- Upon waking up, patient will be asked to urinate and test
- Ask patient to stand up and walk, after 2 hours collect urine sample and test again
- Comapre samples
Urine is contaminated directly or indirectly with albuminous fluids, pus cells, blood, vaginal discharge
Accidental or False or Pseudo
In Accidental or False or Pseudo, what are the constituents which directly or indirectly contaminate urine?
albuminous fluids, pus cells, blood, vaginal discharge
Example of Accidental or False or Pseudo
vaginitis, cystitis
T or F
Pathologic proteinuria are renal diseases and indicated decreased permeability of the glomerular filter
F (indicated increased permeability of the glomerular filter)
Quantification proteinuria
How many g/day is the amount of protein in heavy proteinuria?
> 4g per day
Quantification proteinuria
How many g/day is the amount of protein in moderate proteinuria?
1 to 4 g/day
Quantification proteinuria
How many g/day is the amount of protein in minimal proteinuria?
<1 g/day
Quantification proteinuria
Syndrome associated with heavy proteinuria
Nephrotic syndrome
Yes or No
Is this correlated with nephrotic syndrome?
↑ serum albumin
No, should be ↓ serum albumin
Yes or No
Is this correlated with nephrotic syndrome?
↑ Lipid in blood & urine
Yes
Yes or No
Is this correlated with nephrotic syndrome?
Granular cast
Yes
Yes or No
Is this correlated with nephrotic syndrome?
Glycogen cast
No, should be Fatty cast
Yes or No
Is this correlated with nephrotic syndrome?
Oval fat body
Yes
Quantification proteinuria
What are the conditions correlated with nephrotic syndrome?
- ↓ serum albumin
- ↑ Lipid in blood & urine
- Granular cast
- Fatty cast
- Oval fat body
T or F
Heavy proteinuria means there is high amount of protein in urine and low in blood
T
Quantification proteinuria
Low protein blood is compensated by body through producing of?
lipids (which increase cast)
What are the patterns under Qualitative Proteinuria
Glomerular and Tubular pattern
Qualitative Proteinuria
Glomerular or Tubular Pattern?
more than 3 - 4 g/day
Glomerular Pattern
Qualitative Proteinuria
Glomerular or Tubular Pattern?
(-) charge on GBM
Glomerular Pattern
Qualitative Proteinuria
Glomerular or Tubular Pattern?
Urine (+) large protein
* a2 macroglobulin
* B-lipoprotein
Glomerular Pattern
Qualitative Proteinuria
Glomerular or Tubular Pattern?
Glomerulonephritis
Glomerular Pattern
Qualitative Proteinuria
Glomerular or Tubular Pattern?
1 - 2 g/day
Tubular Pattern
Qualitative Proteinuria
Glomerular or Tubular Pattern?
Urine (+) small proteins
* a1 microglobulin
* b2 microglobulin
Tubular Pattern
Qualitative Proteinuria
Glomerular or Tubular Pattern?
Fanconi, Cystinosis
Tubular Pattern
Qualitative Proteinuria
What are the urine (+) large protein in glomerular pattern?
- a2 macroglobulin
- B-lipoprotein
Qualitative Proteinuria
What are the urine (+) small protein in tubular pattern?
- a1 microglobulin
- b2 microglobulin
Qualitative Proteinuria; T or F
In Glomerular Pattern, the problem is Glomerular basement membrane since it lets even big proteins pass through
True the fire
Qualitative Proteinuria; T or F
In tubular pattern, Glomerular basement membrane is also damaged
F (GBM is intact, tubular is problem)
- due to the overflow of excess level of protein in the circulation
- can be seen with hemoglobin, myoglobin or immunoglobulin loss into the urine
Overflow Proteinuria
Overflow Proteinuria
What is loss into urine which can indicate overflow proteinuria?
hemoglobin, myoglobin or immunoglobulin loss
globin losses heheheh
Causes of Proteinuria
- before plasma reaches the tubules
- Multiple myeloma (Bence Jones Protein) intravascular hemolysis (hemoglobin)
- acute phase reactants
Prerenal
Causes of Proteinuria
- associated with tubes; protein is added as it passes urinary tract
- Glomerular Disorders
- Tubular proteinuria
- Orthostatic proteinuria
- Microalbuminuria
Renal
Causes of Proteinuria
- Lower UTI
- Menstrual contamination
- Vaginal secretions
Post renal
2 common methods in measurement of protein?
- Reagent strip method
- Acid precipitation techniques
2 common methods in measurement of protein
Principle: Protein error of pH indicators
*More sensitive to albumin
*Indicators change in color in the presence of protein
Reagent Strip Method
Measurement of Protein: Reagent Strip Method
pH is at constant at ?
3
Measurement of Protein: Reagent Strip Method
Reagent strip is constant due to what substance which acts as a buffer
Citric acid
Measurement of Protein: Reagent Strip Method
accepts ions from the indicator
Protein (anion)
T or F
Reagent Strip Method is more sensitive to albumin
T
Measurement of Protein: Reagent Strip Method
- contains more amino groups to accept H+ than other proteins
- ideal to observe since this indicates damage in GBR
Albumin
Measurement of Protein: Reagent Strip Method
Indicator used in reagent strip in measurement of protein
tetrabromophenol blue
Measurement of Protein: Reagent Strip Method
Other indicators used in reagent strip in measurement of protein
- Tetrachlorophenol
Tetrabromosulfonaphthalein
Measurement of Protein: Reagent Strip Method
Results:
Negative and Trace value in reagent strip-protein
N/A
Measurement of Protein: Reagent Strip Method
Results:
+
30 mg/dl
Measurement of Protein: Reagent Strip Method
++
100 mg/dl
Measurement of Protein: Reagent Strip Method
+++
300 mg/dl
Measurement of Protein: Reagent Strip Method
++++
2,000 mg/dl
Measurement of Protein: Reagent Strip Method
False positive or False negative?
highly buffered alkaline urine
Sources of error
False positive
acidic strip is override
Measurement of Protein: Reagent Strip Method
False positive or False negative?
high specific gravity
False positive
because specimen is concentrated
Measurement of Protein: Reagent Strip Method
False positive or False negative?
quaternary ammonium compounds
also accepts H ions
False positive
Measurement of Protein: Reagent Strip Method
False positive or False negative?
microalbuminuria
False negative
Measurement of Protein: Reagent Strip Method
False positive or False negative?
proteins other than albumin
False negative
Measurement of Protein: Heat & Acetic acid
Where do you add drops of acid and where do you heat it?
Upper portion
Measurement of Protein: Acid Precipitation Techniques
Principle: precipitation of protein because of heat and acetic acid
* one of the methods under acid precipitation technqiues where you put acid and heat protein
Heat & acetic acid
Measurement of Protein: Heat & Acetic acid
Results:
(-)
no cloudiness or haziness
Measurement of Protein: Heat & Acetic acid
(+1)
diffuse cloud
Measurement of Protein: Heat & Acetic acid
(+2)
granular cloud
Measurement of Protein: Heat & Acetic acid
(+3)
distinct flocculation
Measurement of Protein: Heat & Acetic acid
(+4)
large flocculation dense, sometimes solid
Measurement of Protein
What are the techniques under Acid Precipitation?
- Heat and acetic acid
- 3% Sulfosalicylic acid TEST (Exton’s)
Other mentioned:
- Heller’s Ring or Heller’s Heat or Nitric acid test
- Picric Acid Test
- Robert’s Ring Test
Measurement of Protein: Acid Precipitation Techniques
Also known as Exton’s test
3% Sulfosalicylic acid TEST
Measurement of Protein: 3% Sulfosalicylic acid TEST (Exton’s)
Positive result is?
Formation of precipitation
Measurement of Protein: 3% Sulfosalicylic acid TEST (Exton’s)
Identify what result
Turbidity: No increase in turbidity
Protein range: < 6
Negative
Measurement of Protein: 3% Sulfosalicylic acid TEST (Exton’s)
Identify what result
Turbidity: Noticeable turbidity
Protein range: 6-30
CONGRATS YOU’RE ON THE 200TH CARD
Trace
Measurement of Protein: 3% Sulfosalicylic acid TEST (Exton’s)
Identify what result
Turbidity: Distinct turbidity w/ no granulation
Protein range: 30-100
1+
Measurement of Protein: 3% Sulfosalicylic acid TEST (Exton’s)
Identify what result
Turbidity: Turbidity with granulation; no flocculation
Protein range: 100-200
2+
Measurement of Protein: 3% Sulfosalicylic acid TEST (Exton’s)
Identify what result
Turbidity: Turbidity with granulation & flocculation
Protein range: 200-400
3+
Measurement of Protein: 3% Sulfosalicylic acid TEST (Exton’s)
Identify what result
Turbidity: Clumps of protein
Protein range: > 400
4+
Measurement of Protein: 3% Sulfosalicylic acid TEST (Exton’s)
False positive or False negative?
highly alkaline urine
False negative
it neutralizes since acid is used here
Measurement of Protein: 3% Sulfosalicylic acid TEST (Exton’s)
False positive or False negative?
Iodinated dyes (x-ray contrast media)
False positive
turbid specimen
Measurement of Protein: 3% Sulfosalicylic acid TEST (Exton’s)
False positive or False negative?
Penicillin
False positive
Measurement of Protein: 3% Sulfosalicylic acid TEST (Exton’s)
False positive or False negative?
Salicylate
False positive
Measurement of Protein: 3% Sulfosalicylic acid TEST (Exton’s)
False positive or False negative?
Tolbutamide
False positive
Acid Precipitation Technique: OTHER ACID PRECIPITATION METHODS
Positive result: White opaque ring at the zone of contact
Heller’s Ring or Heller’s Heat or Nitric acid test
Acid Precipitation Technique: OTHER ACID PRECIPITATION METHODS
Positive result: Light cloudiness to a heavy flocculation
Picric Acid Test
Acid Precipitation Technique: OTHER ACID PRECIPITATION METHODS
Positive result: White ring at the point of contact
Robert’s Ring Test
Measurement of Protein
Drug metabolites can cause what result in Reagent strip and Acid precipitation?
Reagent strip: No effect
Acid precipitation: May cause False positive
Measurement of Protein
False positive, False negative, or No effect?
Highly alkaline urine in reagent strip test
False positive
Measurement of Protein
False positive, False negative, or No effect?
Quaternary ammonium compounds in reagent strip
False positive
Measurement of Protein
False positive, False negative, or No effect?
Radiographic contrast media heat and acetic acid tes
False positive
Measurement of Protein
False positive, False negative, or No effect?
High alkaline urine in acid precipitation test
False negative
Measurement of Protein
False positive, False negative, or No effect?
Radiographic contrast media in reagent strip
No effect
What are the methods in quantitative protein?
- Precipitation
- Colorimetric
- Dye binding
Quantitative Protein
- Salicylic acid + Trichloroacetic acid
- Measured by nephelometer or photometer
Precipitation
Quantitative Protein
What reagents does Precipitation use?
Salicylic acid + Trichloroacetic acid
Quantitative Protein
How is Precipitation measured?
nephelometer or photometer
Quantitative Protein
- Trichloroacetic acid + Biuret
- Mixed; solution will be gray
- measured by spectrophotomete
Colorimetric
Quantitative Protein
What reagents does Colorimetric use?
Trichloroacetic acid + Biuret
Quantitative Protein
How is Colorimetric method measured?
spectrophotometer
Quantitative Protein
- Coomassie brilliant blue + Ponceau S
- measured by spectrophotometer
Dye binding
Quantitative Protein
What reagents does dye binding use?
Coomassie brilliant blue + Ponceau S
Quantitative Protein
How is dye binding measured?
spectrophotometer
- Gamma globulin/Light chain immunoglobulin
- Coagulates: between 40 c and 60 c
- Dissolves: 100 c
Associated with:
* Multiple myeloma
* Waldenstrome’s macroglobulinemia
* Malignant lymphomas
Bence Jones Proteinuria
What are the conditions associated with Bence Jones Proteinuria?
- Multiple myeloma
- Waldenstrome’s macroglobulinemia
- Malignant lymphomas
Bence Jones Proteinuria
malignant disorder that results in infiltration of bone marrow by plasma cells
Multiple myeloma
Bence Jones Proteinuria
Bence Jones Proteinuria coagulates at what temp?
between 40 c and 60 c
Bence Jones Proteinuria
Bence Jones Proteinuria dissolves at what temp?
100 c
Bence Jones Proteinuria
What are the methods for Bence Jones Proteinuria
- Heat and HAc Acid test
- Bradshaw
- Toluene Sulfonic Acid (TSA) test
- Electrophoresis
- Immunoglobulin electrophoresis (IFE)
Bence Jones Proteinuria
- indicated by single sharp peak in gamma globulin region
- best method in Bence jones proteinuria
Electrophoresis
- Predictor of clinical nephropathy in insulin dependent diabetes mellitus (diabetic nephropathy)
- Hypertension and cardiovascular risk
- Beginning stages
Microalbuminuria
Presence of microalbuminuria helps detect risk in what diseases?
Hypertension and cardiovascular risk
Microalbuminuria
Specimen for microalbuminuria?
random or first morning urine
Preservation of Microalbuminuria using 24 hour urine specimen?
Refrigeration
T or F
Less than 30 mg is normal in Microalbuminuria using 24 hour urine specimen
T
Amount of albumin present for early kidney disease or Microalbuminuria?
30 to 300 mg
Amount of albumin present for more advanced kidney disease or Macroalbuminuria?
> 300 mg
Albumin Excretion Rate (AER)?
20 to 200 ug/min
What are the methods in Microalbuminuria?
- Micral II Test strip
- Immunodip
- Clinitek microalbumin
- Multistix pro
methods in Microalbuminuria
- Immunologic Test
- Enzyme Immunoassay
- Strip contains gold labeled anti human albumin antibody – enzyme conjugate
- Albumin binds with antibody and move up the strip
- Reacts with the enzyme substrate
Micral Test
methods in Microalbuminuria: Micral Test
What does the strip in micral test contain?
gold labeled anti human albumin antibody (enzyme conjugate)
methods in Microalbuminuria: Micral Test
Antigen:
Antibody:
Antigen: Albumin in urine
Antibody: Reagent
methods in Microalbuminuria: Micral Test
Albumin binds with …….. and move …… the strip
Albumin binds with antibody and move up the strip
methods in Microalbuminuria: Micral Test
The principle used in micral test is reaction with?
reaction with enzyme substrate
methods in Microalbuminuria: Micral Test
sensitivity?
0-10 mg/dl
methods in Microalbuminuria: Micral Test
reagents?
gold-labeled antibody
methods in Microalbuminuria: Micral Test
Enzyme?
2
- B galactosidase
- Chlorophenol red galactoside
methods in Microalbuminuria: Micral Test; T or F
Possible interference in micral test could be due to concentrated urine which results in false positive
F (dilute urine: false positive)
methods in Microalbuminuria: Micral Test
For how long should the strip be allowed to stand
1 min
methods in Microalbuminuria: Micral Test
Positive result?
pink-red
methods in Microalbuminuria: Micral Test
Negative result?
white
methods in Microalbuminuria: Micral Test
This is where the free antibodies reside
part of the micral strip
Zone 1: Conjugate matrix fleece
methods in Microalbuminuria: Micral Test
controls that are fixed
part of the micral strip
Zone 2: Capture Matrix Fleece
methods in Microalbuminuria
- Immunochromatography
- Principle: Immunology
- Albumin in urine binds with blue latex particles coated with anti human albumin antibody
Immunodip
methods in Microalbuminuria: Immunodip
For how long should the strip be left in the urine?
3 mins
methods in Microalbuminuria: Immunodip
Albumin in urine binds with …….. coated with …………
Albumin in urine binds with blue latex particles coated with anti human albumin antibody
methods in Microalbuminuria: Immunodip
migration direction of bound and unbound particle?
migrate up the strip
methods in Microalbuminuria: Immunodip
These particles do not migrate as far as the bound particles
Unbound particles
methods in Microalbuminuria: Immunodip
Positive result?
Darker top band
methods in Microalbuminuria: Immunodip
Negative result?
Darker bottom band
methods in Microalbuminuria: Immunodip
Reagent?
blue latex coated with anti human albumin antibody
methods in Microalbuminuria: Immunodip
Interference in Immunodip happens when urine is dilute which causes false positive results
F (Dilute urine: false negative)
methods in Microalbuminuria
- Albumin:Creatinine ratio
- Principle:Albumin dye binding principle
- More specific & sensitive to albumin
- estimates 24 hr microalbumin excretion
- corrects for overhydration or dehydration in a random specimen
Clinitek Microalbumin
methods in Microalbuminuria: Clinitek Microalbumin
Dye used in Clinitek Microalbumin
diodo dihydoxy dinitrophenyl tetrabrosulphonaphthalein (DIDNTB)
methods in Microalbuminuria: Clinitek Microalbumin
Sensitivity?
8 to 15 mg/dl (80 to 150 mg/dl)
methods in Microalbuminuria: Clinitek Microalbumin
Clinitek microalbumin estimates?
24 hr microalbumin excretion
methods in Microalbuminuria: Clinitek Microalbumin; T or F
Clinitek microalbumin corrects for overhydration or dehydration in a 24 hour specimen
F (corrects for overhydration or dehydration in a random specimen)
methods in Microalbuminuria: Clinitek Microalbumin
produced & excreted at a constant rate
Creatinine
methods in Microalbuminuria: Clinitek Microalbumin
Detection of creatinine in clinitek microalbumin uses what priciple?
Pseudo-peroxide activity of copper-creatinine complexes
methods in Microalbuminuria: Clinitek Microalbumin
These are the involved reactants in clinitek microalbumin
diisopropylbenzene dihydroperoxide (DBDH) and tetramethylbenzidine (TMB)
methods in Microalbuminuria
Multistix pro
What does this indicate:
Protein error of indicators
Protein high
methods in Microalbuminuria
Multistix pro
What does this indicate:
Dye binding method (sulphonapthalein dye)
Protein low
methods in Microalbuminuria
Multistix pro
This substance is based on the Pseudoperoxidase of CuCrea
Creatinine
methods in Microalbuminuria
Multistix or Multistix Pro?
Leukocytes
Multistix and Multistix Pro
methods in Microalbuminuria
Multistix pro
Urobilinogen and bilirubin
Not measured
methods in Microalbuminuria
Multistix or Multistix Pro?
Nitrite
Multistix and Multistix Pro
methods in Microalbuminuria
Multistix or Multistix Pro?
Urobilinogen
Multistix
methods in Microalbuminuria
Multistix or Multistix Pro?
Protein
Multistix and Multistix Pro
methods in Microalbuminuria
Multistix or Multistix Pro?
pH
Multistix and Multistix Pro
methods in Microalbuminuria
Multistix or Multistix Pro?
Blood
Multistix and Multistix Pro
methods in Microalbuminuria
Multistix or Multistix Pro?
Specific Gravity
Multistix and Multistix Pro
methods in Microalbuminuria
Multistix or Multistix Pro?
Ketone
Multistix and Multistix Pro
methods in Microalbuminuria
Multistix or Multistix Pro?
Bilirubin
Multistix
methods in Microalbuminuria
Multistix or Multistix Pro?
Glucose
Multistix and Multistix Pro
methods in Microalbuminuria
Multistix or Multistix Pro?
Protein high
Multistix Pro
methods in Microalbuminuria
Multistix or Multistix Pro?
Protein low
Multistix Pro
methods in Microalbuminuria
Multistix or Multistix Pro?
Creatinine
Multistix Pro
Detectable amount of glucose
Glucosuria (glucose in urine)
Renal threshold of glucose?
160 to 180 mg/dl
2 Types of Glucosuria
- Glucosuria Hyperglycemia associated
- Glucose Renal associated
2 Types of Glucosuria
- Glucose high in blood
- More than 180 mg/dl
- Hormones that cause breakdown of glycogen to glucose
Glucosuria Hyperglycemia associated
2 Types of Glucosuria
What are the conditions associated with Glucosuria Hyperglycemia
- Diabetes mellitus
- Pancreatitis
- Pancreatic Ca
- Acromegaly
- Cushing’s syndrome
- Hyperthyroidism
- Pheochromocytoma
- CNS damage
- Stress
- Gestational diabetes
2 Types of Glucosuria
- Glucose in the blood is normal
- Tubules cannot reabsorb glucose
Glucose Renal associated
2 Types of Glucosuria
What is the main tubule affected in Glucose Renal associated type of glucosuria?
CONGRATS YOU’RE ON THE 200TH CARD
TWERK KA MUNA
Proximal Convoluted Tubule
2 Types of Glucosuria
What are the conditions associated with Glucose Renal
- Fanconi’s syndrome
- Advanced Renal Disease
- Osteomalacia
- Pregnancy
What are the 2 methods used in measuring glucose for glucose renal associated?
Reagent Strip Methods, Copper Reduction Methods
2 methods used in measuring glucose for glucose renal associated
- Specific for glucose only
- Principle: Double sequential enzyme
- Glucose & Peroxidase
Reagent Strip Methods
2 methods used in measuring glucose for glucose renal associated
Principle of reagent strip method?
Double sequential enzyme
2 methods used in measuring glucose for glucose renal associated
ENzymes involved for reagent strip method?
Glucose & Peroxidase
2 methods used in measuring glucose for glucose renal associated
End products of reagent strips?
Oxidized chromogen + Water
2 methods used in measuring glucose for glucose renal associated
Causes of error: Reagent strips
False positive or False negative?
oxidizing cleaning agent (peroxide)
False positive
2 methods used in measuring glucose for glucose renal associated
Causes of error: Reagent strips
False positive or False negative?
Vit C
False negative
2 methods used in measuring glucose for glucose renal associated
False positive or False negative?
Improperly preserved specimen
False negative
2 methods used in measuring glucose for glucose renal associated
False positive or False negative?
High specific gravity
False negative
2 methods used in measuring glucose for glucose renal associated
False positive or False negative?
High ketones
False negative
2 methods used in measuring glucose for glucose renal associated
False positive or False negative?
Low temperatures
False negative
2 methods used in measuring glucose for glucose renal associated
What is the chromogen used for multistix reagent strips?
Potassium Iodide chromogen
2 methods used in measuring glucose for glucose renal associated
What is the negative result in multistix using potasssium iodide chromagen?
Blue
2 methods used in measuring glucose for glucose renal associated
What is the positive result in multistix using potasssium iodide chromagen?
Green-brown in 30 seconds
2 methods used in measuring glucose for glucose renal associated
Chromogen used in chemstrip?
Aminopropyl-carbazole
2 methods used in measuring glucose for glucose renal associated
What is the color transition of chemstrip?
yellow to orange brown
2 methods used in measuring glucose for glucose renal associated
What is the chromogen used in Clinistix?
O toluidine chromogen
2 methods used in measuring glucose for glucose renal associated
What is the color transition of Clinistix?
pink to purple
2 methods used in measuring glucose for glucose renal associated
Reporting for Reagent Strips
Amount present for Negative ?
N/A
2 methods used in measuring glucose for glucose renal associated
Reporting for Reagent Strips
Amount present for Trace ?
100 mg/dl
2 methods used in measuring glucose for glucose renal associated
Reporting for Reagent Strips
Amount present for 1+ ?
250 mg/dl
2 methods used in measuring glucose for glucose renal associated
Reporting for Reagent Strips
Amount present for +2 ?
500 mg/dl
2 methods used in measuring glucose for glucose renal associated
Reporting for Reagent Strips
Amount present for 3+?
1,000 mg/dl
2 methods used in measuring glucose for glucose renal associated
Reporting for Reagent Strips
Amount present for 4+
> 2,000 mg/dl
2 methods used in measuring glucose for glucose renal associated
- Principle: Copper reduction
- Ability of glucose to reduce copper sulfate to cuprous oxide in the presence of alkali & heat
Benedict’s & Clinitest
2 methods used in measuring glucose for glucose renal associated
What are the reagents used in Benedict’s test
- Copper Sulfate
- Na carbonate
- Citric acid
2 methods used in measuring glucose for glucose renal associated
Results for coppper reduction test: Benedict’s & Clinitest
Neg
Blue
2 methods used in measuring glucose for glucose renal associated
Results for coppper reduction test: Benedict’s & Clinitest
Trace
Green w/o precipitate
2 methods used in measuring glucose for glucose renal associated
Results for coppper reduction test: Benedict’s & Clinitest
1+
Yellow w/o precipitate
2 methods used in measuring glucose for glucose renal associated
Results for coppper reduction test: Benedict’s & Clinitest
2+
Yellow green w/ yellow precipitate
2 methods used in measuring glucose for glucose renal associated
Results for coppper reduction test: Benedict’s & Clinitest
3+
Muddy orange w/ yellow precipitate
2 methods used in measuring glucose for glucose renal associated
Results for coppper reduction test: Benedict’s & Clinitest
4+
Orange to red precipitate
T or f
Sucrose tests negative for benedict’s and clinitest
T
2 methods used in measuring glucose for glucose renal associated
- Principle: Copper reduction
- Reagents of benedict’s are in tablet form
Clinitest
2 methods used in measuring glucose for glucose renal associated
CuSO4, NaOH, Sodium carbonate, Sodium citrate
Clinitest
Reagents in clinitest
2 methods used in measuring glucose for glucose renal associated
Citric acid + Na carbonate
Clinitest
Bubbles
2 methods used in measuring glucose for glucose renal associated
NaOH + water
Clinitest
Heat
2 methods used in measuring glucose for glucose renal associated
Negative results for clinitest?
Blue
2 methods used in measuring glucose for glucose renal associated
Positive results for clinitest?
Green to orange/red
2 methods used in measuring glucose for glucose renal associated
If glucose is too high, pass through phenomenon has how much amount of glucose?
Copper reduction test: glucose
> 2g/dl sugar
2 methods used in measuring glucose for glucose renal associated
If glucose is too high, pass through phenomenon goes through what color transition?
Copper reduction test: glucose
Passes through orange to a dark shade of greenish brown
2 methods used in measuring glucose for glucose renal associated
When glucose is high, results pass through orange to a dark shade of greenish brown becasue of what phenomenon?
Copper reduction test: glucose
what specific rection
Re-oxidation of cuprous oxide to cupric oxide
2 methods used in measuring glucose for glucose renal associated
Result: Spotted bluish white tablet
Clinitest
Normal
2 methods used in measuring glucose for glucose renal associated
Result: Dark blue to brown table
Clinitest
Discard specimen
2 methods used in measuring glucose for glucose renal associated
what do you do to the specimen when glucose is too high?
Clinitest
Dilute using two-drop method
2 methods used in measuring glucose for glucose renal associated
Identify what method:
5 drops of urine; 10 drops of water
Clinitest
Five-drop method
2 methods used in measuring glucose for glucose renal associated
Identify what method:
2 drops of urine; 10 drops of water
Clinitest
Two-drop method
2 methods used in measuring glucose for glucose renal associated
Five-drop method results:
Negative?
Clinitest
N/A
Five-drop method results:
Trace
Clinitest
0.25 g/dl
2 methods used in measuring glucose for glucose renal associated
Five-drop method results:
+
Clinitest
0.5 g/dl
2 methods used in measuring glucose for glucose renal associated
Five-drop method results:
++
Clinitest
0.75 g/dl
2 methods used in measuring glucose for glucose renal associated
Five-drop method results:
+++
Clinitest
1.0 g/dl
2 methods used in measuring glucose for glucose renal associated
Five-drop method results:
++++
Clinitest
2.0 g.dl
2 methods used in measuring glucose for glucose renal associated
Two-drop method results:
Negative
Clinitest
N/A
2 methods used in measuring glucose for glucose renal associated
Two-drop method results:
+
Clinitest
1 g/dl
2 methods used in measuring glucose for glucose renal associated
Two-drop method results:
++
Clinitest
2 g/dl
2 methods used in measuring glucose for glucose renal associated
Two-drop method results:
+++
Clinitest
3 g/dl
2 methods used in measuring glucose for glucose renal associated
Two-drop method results:
++++
Clinitest
5 g/dl
Method: Benedict
What is the principle, positive result, and sugar measured?
Principle: copper reduction
Positive result: green, yellow, orange, red
Sugar measured: Glucose and other reducing sugars
Method: Clinitest tablet
What is the principle, positive result, and sugar measured?
Principle: copper reduction
Positive result: green, yellow, orange, red
Sugar measured: Glucose and other reducing sugars
Method: Reagent Strip
What is the principle, positive result, and sugar measured?
Principle: Double sequential enzyme reaction (glucose oxidase)
Positive result: Green to brown
Sugar measured: Glucose only
Method: Reagent Strip
What is the negative result?
blue
Results
Reagent strip: 1+
Benedict’s: -
What is the interpretation?
Glucose (small amount)
Results
Reagent strip: +
Benedict’s: +
What is the interpretation?
glucose & reducing sugars
Results
Reagent strip: -
Benedict’s: +3
What is the interpretation?
non-glucose reducing sugars
Results
Reagent strip: 3+
Benedict’s: +
What is the interpretation?
??????
(strips may not oxidizing agent; benedict reagent is not effective)
Correlation of Reagent Strip and Heat and Acetic acid results:
Reagent strip: +1
Heat &HAc: -
Albumin present
Correlation of Reagent Strip and Heat and Acetic acid results:
Reagent strip: +
Heat &HAc: +
Proteinuria
Correlation of Reagent Strip and Heat and Acetic acid results:
Reagent strip: -
Heat &HAc: +
BJP, Globulins, etc.
Glucose
What is the results on reagent strip and copper reduction?
Reagent strip: +
Copper reduction: +
Reducing sugars
What is the results on reagent strip and copper reduction?
Reagent strip: NE
Copper reduction: +
Homogentisic acid
What is the results on reagent strip and copper reduction?
Reagent strip: NE
Copper reduction: +
Xray dye
What is the results on reagent strip and copper reduction?
Reagent strip: NE
Copper reduction: black
H202 & hypochlorite
What is the results on reagent strip and copper reduction?
Reagent strip: False (-)
Copper reduction: -
Glucose Results
False positive or False Negative?
Ascorbic acid in Clinitest
False positive
Glucose Results
False positive or False Negative?
Oxidizing agents in Reagent strip testing
False positive
Glucose Results
False positive or False Negative?
Ascorbic acid in Reagent strip
False negative
