Diazyme Laboratories
12889 Gregg Court
Poway, CA 92064, USA
Tel: 858-455-4768 / Fax: 858-455-3701
Email: support@diazyme.com
Website: www.diazyme.com
Glycated Serum Protein
Enzymatic Assay
Configuration
The Diazyme Glycated Serum Protein reagent is provided in bulk
and the following kit configuration:
Instrument
Catalog No.
Kit Size
R1: 2 x 20 mL
R2: 2 x 5 mL
Universal
DZ112A-K
Cal: 2 x 1 mL
Reagent Composition
R1: Enzyme/substrate lyophilized powder containing Good’s
buffer, 4-AA, Enzymes and stabilizers
R2: Enzyme/substrate lyophilized powder containing Good’s
buffer, enzymes, TOOS, HRP, and stabilizers
Intended Use
The assay kit is for determination of glycated serum proteins (fructosamine) in human serum.
Clinical Significance1
Fructosamine is formed due to a non-enzymatic Maillard reaction
between glucose and amino acid residues of proteins. In diabetic
patients, elevated blood glucose levels correlate with increased
fructosamine formation. Fructosamine is a medium term indicator
of diabetic control (2-3 weeks).
Assay Principle2
The Diazyme enzymatic assay for glycated serum proteins (GSP)
uses Proteinase K to digest GSP into low molecular weight glycated
protein fragments (GPF), and uses Diazyme’s specific fructosaminaseTM, a microorganism originated amadoriase to catalyze the
oxidative degradation of Amadori product GPF to yield PF or
amino acids, glucosone and H2O2. The H2O2 released is measured
by a colorimetric Trinder end-point reaction. The absorbance at 600
nm is proportional to the concentration of glycated serum proteins
(GSP).
GSP Proteinase K
GPF
H2O2 + TOOS + 4-AAP
Reagent 1 (R1)
Reconstitute one vial Reagent 1 with 20 mL of distilled water. Mix
gently by inversion and then allow to stand for 24 hours at 2-8°C
before use. The reconstituted R1 is stable for 4 weeks at 2-8°C.
Reagent 2 (R2)
Reconstitute one vial Reagent 2 with 5 mL of distilled water. Mix
gently by inversion and then allow to stand at room temperature for
a minimum of 10 minutes before use. The reconstituted R2 is stable
for 4 weeks at 2-8°C.
Calibrator
Reconstitute one vial of calibrator with 1 mL of distilled water.
Mix gently by swirling and then allow to stand at room temperature
for a minimum of 30 minutes before use. The reconstituted calibrator is stable for 4 weeks at 2-8°C.
Reagents from different lots should not be interchanged.
Reagent Stability and Storage
Glycated protein fragments
(GPF)
Fructosaminase
Reagent Preparation
Materials as provided are stable at 2-8°C until the expiration date
noted on the labels. Reconstituted Reagent 1 is stable for 4 weeks
at 2-8°C. Reconstituted Reagent 2 is stable for 4 weeks at 2-8°C.
Reconstituted Calibrator is stable for 4 weeks at 2-8°C.
PF or amino acids + H2O2
Peroxidase
Color + H2O
Specimen Collection and Handling
Use fresh patient serum samples. Serum should be separated from
cells immediately after collection. Samples can be stored at 2-80C
for 2 weeks or at -20°C for up to 4 weeks .
Materials Required But Not Provided
An analyzer capable of dispensing 2 reagents and of measuring
absorbance at 600 nm with temperature control (37°C).
Diazyme Normal and Abnormal controls (Catalog No. DZ112ACON) are recommended.
Diazyme Laboratories
70011 Rev. F
Page 1 of 2
Effective: 2008-01-08
Precautions
1.
2.
3.
4.
Performance Characteristics
Specimens and reagents containing human sourced materials
should be handled as if potentially infectious, using safe
laboratory procedures such as those outlined in Biosafety in
Microbiological and Biomedical Laboratories (HHS Publication Number [CDC] 93-8395).
As with any diagnostic test procedure, results should be interpreted considering all other test results and the clinical
status of the patient.
Avoid swallowing and contact with skin or mucous membranes.
Additional safety information concerning storage and handling of this product is provided within the Material Safety
Data Sheet for this product. To obtain an MSDS, please contact our customer service department at 858-455-4768.
Assay Procedure
R1: 200 μL
Sample: 20 μL o
37 C
0
600 nm
R2: 50 µL
5
A1
10 min
A2
Accuracy
Diazyme vs Randox
65
0.98
0.99
-4.90
N
r
slope
Y intercept
Precision
Intra Assay Precision
Samples
N
Mean (μmol/L)
SD (μmol/L)
CV (%)
Level 1
40
290.10
4.92
1.72
Level 3
40
608.26
10.25
1.75
Total Assay Precision
Samples
Level 1
N
40
290.10
Mean (μmol/L)
8.124
SD (μmol/L)
CV(%)
2.84
Level 3
40
608.26
17.22
2.94
Linearity
Application sheets for use of Diazyme Glycated Serum Protein
Enzymatic Assay on automated clinical chemistry analyzers are
available upon request. Please call 858-455-4768 or email:
support@diazyme.com .
The assay is linear up to 1354 μmol/L.
Interferences
Interference
Ascorbic Acid
Bilirubin
Glucose
Hemoblobin
Uric Acid
Triglyceride
Calibration
GSP calibrator is used for calibration and is supplied in the kit.
Daily calibration is required.
Quality Control
GSP Control Level 1 and Level 3 are recommended to use as daily
quality control sera and can be purchased separately from Diazyme
Laboratories (DZ112A-CON). One control (Level 1 or Level 3)
should be tested at least once a day. The range of acceptable
control limits should be established by individual laboratories.
Concentration
4 mg/dl
20 mg/dl
2400 mg/dl
200 mg/dl
37.5 mg/dl
2000 mg/dl
References
Results
1. Armbuster DA, Fructosamine: Structure, Analysis and Clinical
Usefulness. Clin. Chem. 1987; 33 (12): 2153-2163
Reference Range
2. Kouzuma, T. et al. An enzymatic method for the measurement of
glycated albumin in biological samples. Clin. Chimi. Acta 2002;
324: 61-71
Glycated Serum Protein results are printed out in μmol/L.
Adults (20-60 years) have a reported normal range of 100-285
μmol/L. Each laboratory should establish an expected range with a
set of standards. It is recommended that each laboratory establish
its own reference range to reflect the age, sex, diet and geographical
location of the population.
Limitations
Randox fructosamine calibrators and controls may not be compatible with Diazyme GSP assay.
MDSS
Burckhardtstrasse 1
30163 Hannover, Germany
12889 Gregg Court
Poway, CA 92064, USA
Tel: (858) 455-4754
Fax: (858) 455-4750
Diazyme Laboratories
70011 Rev. F
Page 2 of 2
Effective: 2008-01-08