Bilirubin determination

Question 1

Specimen requirements for bilirubin determination

Answer 1

Avoid light exposure and hemolysis

Question 2

Method of bilirubin determination based on Van den Bergh reaction

Answer 2

Colorimetric-Spectrophotometric

Question 3

Diazo reagents in Evelyn-Malloy method

Answer 3

Diazo A: 0.1% sulfanilic acid in HCl, Diazo B: 0.5% sodium nitrite, Diazo blank: 1.5% HCl

Question 4

Accelerator used in Evelyn-Malloy method

Answer 4

Methanol

Question 5

pH of the reaction in Evelyn-Malloy method

Answer 5

Acidic

Question 6

Color reaction in Evelyn-Malloy method

Answer 6

Red-purple

Question 7

Absorbance maximum for Evelyn-Malloy method

Answer 7

560 nm

Question 8

Reason why Jendrassik-Grof method is better

Answer 8

Alkalinization; not affected by hemolysis

Question 9

Reagents used in Jendrassik-Grof method

Answer 9

Diazo reagents, Sodium acetate, Ascorbic acid (stopping reagent), Alkaline tartrate (Fehling’s reagent)

Question 10

Accelerator used in Jendrassik-Grof method

Answer 10

Caffeine

Question 11

pH of the reaction in Jendrassik-Grof method

Answer 11

Alkaline

Question 12

Color reaction in Jendrassik-Grof method

Answer 12

Blue

Question 13

Absorbance maximum for Jendrassik-Grof method

Answer 13

600 nm

Question 14

Reaction for total bilirubin determination

Answer 14

Bilirubin + Diazo + Accelerator → Azodipyrroles (all fractions)

Question 15

Reaction for direct bilirubin determination

Answer 15

Bilirubin + Diazo → Azodipyrroles (B2 and delta only)

Question 16

Formula for indirect bilirubin calculation

Answer 16

Total bilirubin - Direct bilirubin

Question 17

Principle of direct spectrophotometry (bilirubinometry)

Answer 17

Absorption of light by bilirubin at 480 nm; correction for oxyhemoglobin by measuring absorbance at two wavelengths

Question 18

Appropriate sample type for direct spectrophotometry

Answer 18

Neonatal serum

Question 19

Reason direct spectrophotometry is not done on adult serum

Answer 19

Presence of carotenoid pigments

Question 20

OD 480 principle application

Answer 20

Hemolytic Disease of the Fetus and Newborn (HDFN)

Question 21

Principle of enzymatic bilirubin determination

Answer 21

Oxidation of bilirubin to biliverdin by bilirubin oxidase, with absorbance decrease measured

Question 22

Reference value for B1

Answer 22

0.2-0.8 mg/dL

Question 23

Reference value for B2

Answer 23

0-0.2 mg/dL

Question 24

Source of B2 and its normal destination

Answer 24

Secreted from canaliculi; should be in the intestine, not in the blood

Question 25

Reference value for total bilirubin

Answer 25

0.2-1.0 mg/dL

Question 26

Bilirubin level indicating overt jaundice

Answer 26

TB > 3.0 mg/dL

Question 27

Panic value for kernicterus

Answer 27

TB ≥ 15 mg/dL