clinical urinalysis and body fluids review, part 1

Clinical Urinalysis and

Body Fluids Review, part 1

Austin Community CollegeMedical Laboratory TechnologyClinical II Spring 09

Body Fluids other than urine

What general purposes do the body fluids serve? Nutrition Waste removal Lubrication Cushioning / protection

Body Fluids Types

CSF Serous (transudate versus exudate)

Peritoneal Pleural Pericardial

Synovial Seminal Amniotic Sweat , gastric, feces, etc.

Body Fluids Testing

Gross and hematological Cell counts Differential

Chemistry TP Glucose Enzymes?

Amylase, Lipase and LDH Microbiology – cultures Serology? Cytology ?

Body Fluids

CSF Why evaluate?

Body Fluids

CSF Why evaluate?

Diagnose meningitis Evaluate for intracranial hemorrhage Diagnose malignancies, leukemia Investigate central nervous system disorders

Body Fluids

CSF What normal term(s) are used to describe

the color and clarity of CSF specimens? Identify terms used for ‘abnormal’ colors

and clarity. What is that ‘special’ color term unique to

CSF? How do you differentiate between traumatic

tap and cerebral hemorrhage?

Body Fluids

CSF Reference ranges / normal or expected

values

Body Fluids

What is the term used when there is increased numbers of cells in CSF?

Pleocytosis Can be prefixed as

Neutrophilic pleocytosis Lymphocytic pleocytosis, etc.

Body Fluids

Why evaluate serous and synovial? To determine the reason for increased

production

What’s the term that is used to indicate increased production of these fluids?

Body Fluids

What’s the term ?

Effusion – an increase in the serous fluid due to some disruption in production and/ re-absorption processes.

Seminal fluid

In what tissue are sperm produced?

Four reasons for analyzing seminal fluids

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Seminal fluid

In what tissue is sperm produced? Spermatozoa - produced in the testes, mature in

the epididymis. Four reasons for analyzing seminal fluids

Infertility issues Post- vasectomy Forensic analysis Sperm donors

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Body Fluids other than Urine

What are the collection requirements?

Normal appearance?

Expected lab values? Cell count Motility Morphology

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What is the formula for calculating body fluid cells counted in a hemacytometer?

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What is the basic formula?

What additional calculation is needed for sperm counts and why?

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ave . # ce l ls counted x d ilution# squares counted x vo lume of each square


What additional calculation is needed and why? the standard calculation provides results as per

microliter (uL) And sperm normal value is per milliliter (mL) So you have to take the results and multiply X

1000.

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What seminal fluid substances can be used for forensic and rape evaluations?

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What seminal fluid substances can be used for forensic and rape evaluations? Acid phosphatase enzyme DNA UV light ABO /HLA

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Amniotic Fluid Of what is it composed? What is the name of the collection

procedure? What purpose does it serve? Why is it sometimes analyzed?


Amniotic Fluid Indications for analysis

Chromosomal abnormalities, such as Down’s syndrome

Metabolic disorders, such as Tay Sachs Neural tube defects – such as spinal bifida or

an encephalic Determination of extent of HDN Others - gestational age, fetal maturity, etc.


Why do we sometimes analyze sweat? What is the name of that procedure? What are normal values?


Why do we analyze gastric fluids? How are the specimens collected? What lab procedures are performed?

Body Fluids other than Urine Indications for testing

Drug analysis Peptic ulcer evaluation

Specimen collection Nasal or oral intubation Fasting and avoid swallowing saliva

Laboratory procedures Gastric acidy (acid <7.0)

Peptic ulcers, diagnosis and treatment Zollinger-Ellison Syndrome – tumor of pancreas

causing gastric over-secretion Decreased gastric secretion Anacidity – inability to produce acid

Drug screening


Feces What is it and why do we care? What is the significance of different stool

colors?


What test is performed as a screen for colorectal cancer?


What patient preparation is needed for this test to be most useful?


What other tests are done on fecal samples?

Cells of the Body Fluids

What cells can be found in body fluids? Cells seen in the peripheral blood, but they

sure don’t look so good in the BFs.

Cells of the Body Fluids (other than

Urine) More often they look like these:

Lymphocytes and monocytes Cerebrospinal Fluid (CSF)

Cells of the Body Fluids (other than

Urine)

Lymphocytes and eosinophils – CSF Patient had shunt


Lymphocytes, macrophages and basophil

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Cells of the Body

Other cells Ependymal cells - on left Mesothelial cells - on right

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Cells of the Body Fluids Then there are the inclusions, and the ‘new’ names


And the ‘other’ cells, ‘unclassified’ – those referred to the cytologist / pathologist.

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Intracellular inclusions: The yeast

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The yeast - Cryptococcus in CSF

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Bacteria

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For synovial fluid Crystals

Calcium pyrophosphate Monosodium urate

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Serous and Synovial Fluids

LE cells –

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Clinical Urinalysis and

Body Fluids Review, part 2

Austin Community CollegeMedical Laboratory TechnologyClinical II Spring 09

Chemical Exam of Urine


Reagent strip manufactures Bayer Corporation- Diagnostics Division

(formerly Ames) produces Multistix

Boehringer-Mannheim Corporation which produces Chemstrip

Behring Diagnostics which produces Rapignost


Recall normal dipstick procedure

What are sources of error?


Sources of error (and preventions) Testing cold specimens

would result in a slowing down of reactions; test specimens when fresh or bring them to RT before testing

Inadequate mixing of specimen could result in false reduced or negative

reactions to blood and leukocyte tests mix specimens well before dipping

Over-dipping of reagent strip will result in leaching of reagents out of pads;

briefly, but completely dip the reagent strip into the urine


Inadequate blotting and Failure to keep strip horizontal will result in over-run or mixing of reagents

between the different reaction pads; blot excess urine off the strip and keep strip horizontal. If dipping from the tube, can run the side of the strip along the rim to remove excess urine.

Improper timing of tests over development of reagent pad colors leading to

falsely increased results; follow manufacturer’s recommendations


Inadequate light misinterpretation of results; use good lighting

Mis-using color chart misinterpretation of results; hold strip just

over color chart and match colors as close as possible, consider use of back-up tests, if needed, especially if urine’s color masks reaction colors.


Handling and Storage Keep strips in original container, stored at

RT Protect from moisture and volatile fumes Use before expiration date Do not touch reagent pad areas


Quality Control - use appropriate, commercially prepared positive and negative controls. Use commercially prepared pos and neg

controls, at least once per 24 hours, and anytime a new bottle is opened, or question of validity of results.

Readings should agree with published results ± one color block.

Urine Glucose Testing

Normal : no glucose detected Clinical significance of abnormal results

(Glucosuria) Sensitivity

Approximately 50-100 mg (compared to Clinitest’s 250) SO- Can have a positive dipstick but a neg Clinitest

Specificity - is specific for glucose only. not affected by other sugars or reducing

substances.

Urine Glucose Testing

Interfering substances

High specific gravity and high pH may depress color.

Ascorbic acid-false neg Bleach or peroxide may give false positive

Urine Bilirubin Testing

Normal : no bilirubin detected

Clinical significance of abnormal results (Bilirubinuria) Jaundice types


Testing method Urine dipsticks for bilirubin – a diazo

reaction Impregnated with stabilized diazotized 2,4

dichloraniline Color goes from buff to brown also shades of

pink – violet

If urine is strongly colored, look for change in pad color after dipping.

Use Ictotest for backup.


Interfering substances

Medication metabolites, pigments and indican may obscure readings

False negatives due to aged specimens, especially those exposed to light and oxidation.

Urine Ketone Testing

Ketone Bodies Origin - not normally present

Products of fat catabolism - breakdown of fat into CO2 and H2O


Ketone Bodies What are the 3 ketone bodies?

What effect do they have on the body?

What is the testing methodology?


Acetone 2%. -Acetone is volatile and excreted primarily through

the lungs

Diacetic Acid (Acetoacetic) the first formed, 20 % of the total the form detected by most ketone test procedures

Beta hydroxybutyric Acid majority formed, but not detected by routine tests. Only Hart’s test, an old ‘wet chemical’ test will detect

this one.


Clinical effects Metabolic acidosis

Lowering of blood and urine pH Brain toxicity


Testing Most use nitroprusside

Detects diacetic acid and a small amount of acetone, but does not detect β-hydroxybutyric acid.

Produces purple color. Can be used on urine or blood.

Urine Specific Gravity Testing

The specific gravity is a measure of the weight of urine compared to an equal amount of water. Water has an assigned value: 1.000

Specific gravity is related / proportional to urine osmolality; both are measures of concentration.


Specific gravity between 1.002 and 1.035 on a random sample is normal if kidney function is normal.


Dilute urine will have values less than 1.010. Fixed specific gravity = 1.010; isothenuria Diabetes insipidus End-stage renal disease

And concentrated urine will have values usually over 1.020. Usually due to dehydration and can be seen

in well population as well as sick.

Urine Specific Gravity Testing Increased urine specific gravity may be seen in:

Dehydration Diarrhea Excessive sweating Glucosuria Heart failure (related to decreased blood flow

to the kidneys) Renal arterial stenosis Syndrome of inappropriate antidiuretic

hormone secretion (SIADH) Vomiting Water restriction


Decreased urine specific gravity may indicate / be seen in: Excessive fluid intake Diabetes insipidus – central or

nephrogenic Renal failure (that is, loss of ability to

reabsorb water) Pyelonephritis


Specific gravity > 1.035 (refractometer) Could have very high glucose levels Could contain radiographic dye


Testing Polyelectrolytes , pH indicator (bromthymol

blue measures the pH change) and alkaline buffer


Interfering substances False elevation of results may be seen in

samples with increased protein concentration.

Some reports of reduced specific gravity results on alkaline specimens.

Lipids may also effect results

Urine Blood Testing

Normally not found in urine Hemoglobinuria – free hemoglobin in urine

Circulating free hemoglobin normally picked up by haptoglobin preventing loss in urine

When serum levels of hemoglobin > 100 mg/dL threshold is exceeded

Hematuria – RBCs in the urine Trauma / irritation of renal organs

Urine Blood Testing

Urine Blood Testing

Testing dipstick reaction

HGB H2O2peroxidase OxygenHGB H2O2peroxidase Oxygen

Oxygen + Gum guaiac, benzidine or orthotolidine green or blue

oxidation products

Urine Blood Testing

‘Blood’ test detects Free Hemoglobin RBCs – get lysed on the pad and their

hemoglobin reacts Myoglobin – muscle hemoglobin

Principle based on the peroxidase-like activity of the heme portion of the molecule

Urine Blood Testing

Sensitivity – can detect at levels of 5-10 cells/uL

Interfering substances Ascorbic acid Nitrates Oxidizing agents (ie bleach) Contaminate blood (menstrual)

Urine pH Testing

Normal: kidneys capable of 4.5 – 8.0 Factors effecting pH

Diet – general and specific foods Time of day Metabolic disorders Drugs / medications

Dipstick capable: 4.5 – 9.0

Urine pH Testing

Test method Dipstick indicators – methyl red and

bromthymol blue Range 4.5-9.0

Caution – other chemicals on dipstick can effect pH reading

Urine Protein Testing

Normally not found in measurable amounts on dipstick (<150 mg/dL /day) Permeability of glomerulus

Damage to glomerular capillaries Changes in glom blood flow

Albumin excretions may be increased temporarily due to exercise, UTI, and acute illness with fever.

Dipstick results of > 1+ (30mg/dL) would equal to approximately 500 mg/dL (clinical proteinuria)


Only albumin detectable by dipstick Sensitivity (approximately15-30 ml/dL)


New testing for microalbumin and creatinine Results:

Protein 20-200 mg/dL (30-300 mg/dL /24 hr) Creatinine 10-300 mg/dL

Albumin/Creatinine ratio Normally albumin in the urine is less than 30 mg/

gram creatinine


Principle - Protein error of indicators at fixed pH, certain indicators show one color in

the presence of protein and another in absence of protein - the “error” of the indicator.

Indicator – tetrabromphenol blue - can be hard to read at the trace end

Citrate Buffer – maintains pH 3 -quite acid


Sources of error Sensitive only to albumin Urine with strong / unusual color makes

reading difficult Highly alkaline or buffered urine will

neutralize acid buffer and lead to increased erroneous results.

Urine container contamination would interfere


Urine back up test 3% sulfosalicylic acid

Added to the supernatant to detect any kind of protein. Urine will turn cloudy if protein is present.

Urine Urobilinogen Testing

Normally found in small amounts, especially in early afternoon

Increased amounts may indicate liver disease or be seen as result of hemolytic disorders

Decreased amounts: If intestinal bacteria destroyed Liver doesn’t conjugate bilirubin Biliary obstruction – failure of bilirubin to

reach small intestine

Urine Urobilinogen Testing

Test principle based on Ehrilich’s reaction

Para-dimethylaminobenzaldehyde = Ehrlich's reagent.

Must protect specimen from light and test immediately

Urine Nitrate Testing Nitrate

Detects presence of certain types of bacteria screening for presence of UTI. Certain species of bacteria convert nitrate

(normal constituent of urine) to nitrite Escherichia - most common cause of UTI Klebsiella Proteus Pseudomonas Enterobacter Citrobacter

Urine Nitrate Testing

Aromatic amine in reagent strip reacts with nitrite; producing a diazonium salt

The diazonium salt reacts with sulfanilic acid and acetic acid to produce a pink azo dye


Limitations reported as positive or negative Not all UTI causing bacteria convert nitrate to

nitrite Haemophilus Staphylococcus Streptococcus


Fresh first morning specimen is preferred - besides being the most concentrated specimen, the urine has been in the bladder longer, allowing bacteria time and opportunity to convert the nitrates to nitrites.

Urine Leukocyte Testing

Leukocyte esterase testing is another test used as a means of screening for urinary tract infection.

Does not measure concentration of leukocytes

Will detect presence of lysed leukocytes as well as intact WBCs


test principle: Leukocyte esterase, an enzyme present in

granulocytes, hydrolyzes indoxylcarbonic acid esterase to produce indoxyl, which reacts with a diazonium salt to create a purple color usually in 2 min.


Reaction interference False positives - oxidizing detergents False negatives - greatly increased glucose,

protein, or specific gravity- increased sp gr could cause WBC to crenate preventing their releasing their esterase, So it is possible for the dipstick to be negative when there are WBCs present.

Microscopic Sediment

A number of slides with microscopic elements

RBC, WBC, yeast

Microscopic Sediment – Epithelial Cells Squamous epithelial cells (stained with

Sternheimer-Malbin)

Microscopic Sediment – Epithelial Cells

Transitional epithelial cells Spherical, polyhedral and caudate are terms

describing shapes. All have distinct centrally located nuclei. Sometimes called bladder cells, may be more often

found in elderly. Can be found as fragments or as reactive.

Microscopic Sediment – Epithelial Cells RTEs; 250x magnification Also WBC and RBCs

Microscopic Sediment – Casts

Microscopic Sediment – Casts

RBC cast


A number of slides with microscopic elements CRYSTALS, Uric acid

Microscopic Sediment – Crystals

Calcium oxalate – envelope; may be dumbbell shaped. Usually appear as a square with a retractile cross


A number of slides with microscopic elements Alkaline


A number of slides with microscopic elements

The rarely ever seen, abnormal crystals Cystine Tyrosine Leucine Bilirubin (occasionally seen in premies) Cholesterol

Microscopic Sediment A number of slides with microscopic

elements Mucous threads Bacteria, yeast, Trichomonas sperm Lots of artifacts

Microscopic

What are oval fat bodies?

How can you (quickly, cheaply) provide tentative proof they contain fat?

Microscopic Sediment – Miscellaneous

Oval Fat Bodies This frame shows an oval fat body ("B") next to several

transitional epithelial cells ("A"). Note the drops of lipids that appear to be contained within the cell.” – U of IA

clinical urinalysis and body fluids review, part 1

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