Clinical Pathology Laboratory Activity EMS 2012

• Blood Glucose Measurement• Kketone and Glucose Urine Exam

• Blood Gas Analysis• Cholesterol Examination• Cortisol Measurement

Factors influencing laboratory results

Pre analytic

Analytic

Post analytic

Glucose Measurements

SPECIMEN CONSIDERATIONS

Whole blood Plasma Serum Pleural fluid CSF Urine

Most measurements by enzymatic methods:

1. glucose dehydrogenase, 2. glucose oxidase, 3. hexokinase

Glucose Measurements

1. Glucose Dehydrogenase

2. Glucose Oxidase,

3. Hexokinase

(Henry JB. Clinical Diagnosis and Management by Laboratory Methods. 2011)

• These reactions produce an electrical current that is proportional to the initial glucose concentration, or a product that measured spectrophotometrically is proportional to the initial glucose concentration.

• The assays can be initial rate-of-change assays, where the velocity of the reaction is dependent on the initial glucose, or end-point assays.

Glucose Measurements

Spectrophotometer

POCT (point of care testing)/ Blood Glucose Home Monitoring/ Self Glucose Home Monitoring

Glucosemeter

Spectrophotometer

POCT (point of care testing)/ Blood Glucose Home Monitoring/

Self Glucose Home Monitoring

Method of POCT (point of care testing)/ Blood Glucose Home Monitoring/

Self Glucose Home Monitoring

Biosensor

Reflektansmeter

Biosensor Method

Biosensor Method

Reflektansmeter/ Amperometer Method

A wide variety of devices are available forhome measurements:

• avoiding operator errors• Calibration• Errors that may contribute to inaccurate readings in certain

devices include the application of :- an insufficient volume of blood- milking the finger to acquire sufficient blood- the use of outdated test strip- environmental factors (humidity, heat, altitude)- the use of a malfunctioning meter- the use of a dirty meter - hypertriglyceridemia- hypotension- measurements outside of the hematocrit or temperature range

Influecing Factors

• Influenced by high levels of salicylate, acetaminophen, levodopa, uric acid, bilirubin,lipids, or low oxygen levels, and others are altered by touching the reaction area.

• Vit C, lactose, manose, galactose, xylose .

Reference Value

Ketone and Glucose Urine Exam

Dipstick Urine Test/ Strip

Urine test strip:Rapid, easy, specific,cheap

SPECIFIC GRAVITY

PROTEIN

KETOBODY

UROBILINOGEN

BLOOD

PLASTIK ROD

NYLON COVER

NITRITE

GLUCOSE

TEST FIELD(PAPER CONTAIN REAGENT)

FILTER PAPER

DIPSTICK TEST

Dipstick urine test

1. Specific gravity2. pH3. Leukocytes4. Nitrates5. Protein6. Glucose7. Ketones8. Urobilinogen9. Bilirubin10. Blood

Glucose (negative)

Glucose is normally present in glomerular filtrate, but it is reabsorbed by the proximal tubule.

D-Glucose + O2 δ-D gluconolactone + H2O2

GOD

H2O2 + 0-tolidine H2O + colourPOD

Ketones (negative)

in healthy individuals, ketone bodies are formed in the liver and are completely metabolized so that only negligible amounts appear in the urine

sodium nitroprusside and glycerin + acetoacetate and acetone

alkaline medium violet dye complex.

URINALYSIS STEPS

PRE-ANALYTIC : patient preparation, samples collection, samples handling, labelling, refrigeration, preservatives of urine specimens.

ANALYTIC: principle of procedures, measurements, interpretation, conventional & rapid and sophisticated

POST-ANALYTIC: recording, reporting, use of units : conventional unit and international unit

QUALITY CONTROL (QC):calibration, control solution to get good and reliable results

Pre-Analytic1. Specimen collection- Requisition form must accompany with specimen

delivered to the lab; the form include : patient’s name, I.D number, date and time of collection and additional information : age, location, physician’s name, type of specimen/method, interfering medication and clinical information

- Container clean, dry, leak proof, disposable - All specimens must be properly labeled must be attach

to the container, not to the lid, should not become detached if the container is refrigerated

- The information on form requisition match with the inform on the label

Labelling urine specimen

Name (min 2 initial, ex: Deni Darmawaty)

Sex : female

Date (day, month, year)

Time collection:

Adress and telp number :

Pre-AnalyticTwo Type of Specimen Based on Time :

- Random specimen- First morning/fasting specimen- 2-hours post prandial specimen- Timed urine (12-hours, 24-hours specimen)

Based on method :- Midstream Clean-Catch specimen- Catheterized specimen- Suprapubic aspiration

Type of Urine Specimens Purpose

Random Routine screening

First morning Routine screeningPregnancy testsOrthostatic protein

Fasting (second morning) Diabetic screening/monitoring

2-h postprandial Diabetic monitoring

Glucose tolerance test Accompaniment to blood samples in glucose tolerance test

24-h (or timed) Quantitative chemical tests

Catheterized Bacterial culture

Midstream Clean-catch Routine screeningBacterial culture

Suprapubic aspiration Bladder urine for bacterial cultureCytology

• The first morning urine:

Collected upon rising, it represents the urine over

approximately an 8 hour period

• Ad random urine:

Collected any time

• The 2-hour postprandial urine:

Collected 2 hour following the meal ( for urine glucose)

• The 24-hour urine:

A pooling of all urine excreted by the patient over a 24 hour period

(for protein, uric acid, calsium quantitation, etc)

• Midstream urine:

The middle portion of a single urination

Pre-Analytic

3. Specimen HandlingFolowing collection should be delivered to the lab promptly and tested within 1-2 hoursIf it can’t delivered must be refrigerated or add with chemical preservative

LIMITATION OF THE METHOD AND INTERFERING FACTORS:

• Highly pigmented or large amounts of levodopa metabolites in urine may cause weak positive result

• High Specific Gravity and low pH urine and PSP (phenolsulfophthalein) may cause false positive result

• High-protein, carbohydrate-free, high-fat diets may result in ketonuria (false posi- tive)

• Medications : phenazopyridine , ascorbic acid, ether, insulin, isopropyl alcohol, metformin, isoniazide, isopropanol, paraldehyde, valproic acid, and bromsulfoph- thalein also can cause false positive results

EXPECTED VALUE

• In starvation diets or in other instances of abnormal carbohydrate metabolism, ketone appear in urine in excessively large amounts before serum ketone are elevated.

• This test detected 5 mg/dL of aceto-acetic acid; 70 mg/dL acetone; but more specific for aceto-acetic acid.

Lipoprotein structure