LIQUIZYMe

BILIRUBIN KIT
(Mod. Jendrassik and Grof's Method)
INTENDED USE: BEACONN
Bilirubin is used for the detemination of Direct and Total
Bilirubin in serum.

PROCEDURE:
SUMMARY: Pipette into clean dry test tubes labeled as Blank (B) and Test(T)
Bilirubin is mainly formed from the heme portion of agedor
fom a Total Billrubin
damaged RBC'S. t then combines with albumin to
complex, which is water-soluble. This
not is referred to as indirect Additilon sequence (B) (T)
or unconjugated Bilirubin. In the liver this Bilirubin complex is
Total Blirublin Reagent 10 m 10n
combined with glucuronic acid into a water-soluble conjugate
This is referred to as conjugated or direct Blirubin. Elevated 50
Total Nitrite Reagent
levels of bilirubin are found in liver diseases (Hepatitis, cirrhosis)
excessive heemolysisi destruction of RBC (hemolytic jaundice) Sample 100 100
obstruction of the biliery tract (obstructive jaundice) and in drug
induced reections. The differentation between the direct bilirubin Mix well and incubate at R.T. for exactly 10 minutes.
and indirect blirubin is important in diagnosing the cause of Measure the absorbance of the Test Samples
Hyperbilirubinemia.
(Abs. T) immediately against their respective Blanks
PRINCIPLE:
Bliubin reacts with diazotised sulphanilic acid to fomm a coloured Direct Bilirubin
azobilirubin compound. The unconjugated bilinubin couples with Addition sequence (B) (T)
the sulphanilic acid in the presence of a caffein-benzoate Direct Blirubin Reagent 1.0 m 1.0 ml
accelerator. The intensity ofthe colour fomed is directly
proporional to the amount of bilirubin present in the sample. Direct Nitrite Reagent 50

REACTION: Sample 100 100

Bilirubin Diazolized Sulphanilic acid Azobilirubin

Compound
Mix well and incubate at R.T. for exactly 5 minutes.
Measure the absorbance of the Test Samples
(Abs. T) immediately against their respective Blanks
cONTENTS:
CALCULATION:
Reagent 1: Direct Bilirubin Reagent
Reagent 2 : Direct Nitrite Reagent Bilirubin mg/dl(Total or Direct)=Abs. of Test-Abs of Blank x Factor
"Factor = 13
Reagent 3: Total Blirubin Reagent
Reagent 4 : Total Nitrite Reagent LINEARITY:
Reagent 5 : Artficial Standard (10.0 mg/dl)

This procedure is linear upto 20 mg/dl. If the values exceed this

limit, dilute the sample with normal saline (NaCi 0 9% Jand
MATERIALS REQUIRED BUT NOT PROVIDED: repeat the assay Multiply result by dilution factor
Clean&Dry Glassware
Laboratory Glass Pipettes or Micropipettes&Tips.
Bio-Chemistry Analyzer NORMALVALUE
Serum
SAMPLES: Total Bilirubin upto 10 mg/d
Serum. Bliubin is reported to be stablei the sample for 4 days Direct Blirubin upto 0 2 mg/d
at2-8'C protected from light, as it is photosensitive Each Laboratory should estatblish it's own normal range
representing its patient population
PREPARATONOF REAGENT&STABILITY
All reagents are stable at room
temperature (22-30'C) till expiry
QUALITY CONTROL
For accuracy it is necessary to run known controls with every
date mentioned on the label
assay
GENERAL SYSTEM PARAMETERS: LIMITATION&PRECAUTIONS:
Reaction type End point 1.It is important to ensure through mixing of Blirubin reagent
Wave length 546nm (530-550nm) and nitrite reagent before assay
2. Bllrubin levels may be reduced if the sample are exposed to
Temperature R.1
Incubation 10 min. (Total Bilirubin) ight
6 min. (Direct Blirubin) 3 The limit of detection for Direct Bilirubin & Total Bilirubin is
0 03 mg/d
Reagent volume 1.050 ml
Sample volume 100
Standard Concentration 10.0 mg/d
Zero setting Deionised water
Light path 1.0 cm
Factor 13

BEACON DIAGNOSTICs PVT. LTD 424, NEW GIDC, KABILPORE, NAVSARI -.396 424. INDIA
Interferences:
For diagnostic purposes, results should always be assessed in conjunction
with the patient's medical the
history, clinical examination and other findings.

Note:
the artificial standard (S) may be
In case the exact wavelength is not available
artificlal standard against distlled water
used. Measure the absorbance of the
future calculations by divlding
with the appropriate filter and keep the same for
Discard the artificial standard after
the Abs T with the Abs. Of the std. X 10.
use and the samples
In of neonates where the sample quantity is a limitation,
case
of the sample may
have highbilirubin (above 3 mg/dl), only 0.05m/0.02ml
calculation factor in this case would be
used for bilirubin estimation. The
24.9/60.5 respectively instead of 13. In case
of using the standard the value of
instead of 10 mg/dl. The
the same would be 19.1/46.5 mg/dl respectively
automated analyzers.
reagent may be used in several

BIBLIOGRAPHY
1. H. Gowen lock, Maurice Bell et. al. Practical
varley, A. H. 1024.
clinical
Biochemistry. 5th edi. P. 1022 to
2. Jendrassik L. & Grof P. Biochem 2.297, 81 (1938)

4 Reagent 5
CODE NO. PACK SZE Reagent 1 Reagent 2 Reagent 3 Reagent
2x 100 ml 1x 100 ml 1x5.0 ml 1x 100 ml 1x 5.0 ml 1 x 5.0 ml
s27

CE VD C
EC REP And iso 9001:2015J oDSCIC