Chemical Examination of Urine Part III: Ketones, Blood, Bilirubin & Urobilinogen Ricki Otten...

Chemical Examination of Urine

Part III:Ketones, Blood, Bilirubin &

Urobilinogen

Ricki Otten MT(ASCP)SC

uotten@unmc.edu

2

Ketones: Purpose

• Ketones are intermediary products of fat metabolism

3

Ketones

• Three ketone bodies– Acetone 2%– Acetoacetic acid 20%– Beta-hydroxybutyric acid 78%

• Characteristic ‘fruity breath’ ~ acetone

4

Ketones: Normal

• Normal: negative

• Abnormal:– Inability to utilize carbohydrates– Excessive loss of carbohydrates– Inadequate intake of carbohydrates

5

Ketones: Methods

• Reagent strip

• Acetest: tablet test

6

Ketones: Method

• Glycine: also measures acetone– Reagent strip: check package insert– Acetest tablets: contain glycine

7

Ketones• Reagent strip

– Sensitivity: 5-10 mg/dl– Specificity: acetoacetic acid and/or acetone

• False positive: highly pigmented urine• False negative: improper specimen handling

• Acetest– Specificity: acetoacetic acid and acetone

• False positive: highly pigmented urien• False negative: improper specimen handling

8

Blood: Purpose

• Blood in urine indicates pathology

• Two forms found in urine– Intact RBC– Hemolyzed RBC

9

Blood: Terms

• Hematuria

• Hemoglobinuria

• Myoglobinuria

All will give a positive blood reaction

10

Blood: Reagent strip

• Test can detect hemolyzed RBC

• Heme moiety imparts peroxidase activity and catalyzes the reaction

11

Blood

• Sensitivity

• Specificity– Intact RBC– Hemolyzed RBC (hemoglobin)– Myoglobin

– False positives: myoglobin, oxidizing agents– False negatives: ascorbic acid

12

Blood:

• Correlate reagent strip results– Microscopic findings– Color and clarity

13

Bilirubin and Urobilinogen

• Bilirubin in urine is always pathologic:

liver disease

• Urobilinogen in urine: normal to have a small amount:

0.2 – 1.0 mg/dl

14

Three mechanisms

• Pre-hepatic: liver is healthy

• Hepatic: liver disease

• Post-hepatic: liver is healthy, obstruction indicated

15

Bilirubin: Methods

• Reagent strip

• Ictotest: tablet test

• Foam test

16

Bilirubin: Methods

• Reagent strip

• Ictotest: tablet test

• Same reaction

• Same specificity: conjugated bilirubin– False positive: urine color– False negative: low concentration, ascorbic

acid, improper specimen handling

17

Bilirubin: Methods

• Reagent strip

• Ictotest: tablet test

• Sensitivity differs

Reagent strip: ~0.5 mg/dl

Ictotest: 0.05 – 0.1 mg/dl

18

Bilirubin: Methods

• Possible to have a negative reagent strip test and positive ictotest– Difference in sensitivity levels

• Always perform Ictotest when– Urine bilirubin test specifically ordered– Urine appearance is amber: even if bilirubin

reagent strip test is negative– Positive reagent strip test

19

Bilirubin: Foam Test

• Shake urine and observe resulting foam

• Yellow foam = bilirubin

20

Urobilinogen: Methods

• Reagent strip test– Two reactions dependent upon manufacturer

• Para-dimethylaminobenzaldehyde• Diazonium salt

– Cannot determine absence of UBG

• Watson-Schwartz assay

21

Urobilinogen: Methods• Para-dimethylaminobenzaldehyde

– Sensitivity: 0.2 mg/dl– Specificity:

• False positive: any ‘Ehrlich reactive compound’; color masking; urine at body temp

• False negative: improper specimen handling

• Diazonium salt– Sensitivity: 0.4 mg/dl– Specificity: reacts only with UBG

• False positive: color masking• False negative: improper specimen handling

22

Urobilinogen: Watson Schwartz

• Classic method used to differentiate

urobilinogen from porphobilinogen using a

differential extraction method

• Para-dimethylaminobenzaldehyde