EC E N Z Y Cholesterol Oxidase M ES (Recombinant) · MES BICINE MOPS CHES KPO4 Na ACETATE PHOSPHATE...
Transcript of EC E N Z Y Cholesterol Oxidase M ES (Recombinant) · MES BICINE MOPS CHES KPO4 Na ACETATE PHOSPHATE...
O R I G I N Microbial
� S P E C I F I C A T I O N SAppearance Yellow free flowing powderActivity >32 U/mg powder at 37°CSpecific Activity >45 U/mg protein at 37°CContaminants Catalase <1%
Glucose Oxidase <0.01%Uricase <0.01%
� U N I T D E F I N I T I O NOne unit of activity is defined as the amount of enzyme that will catalyse the oxidation of 1.0micromole of cholesterol per minute at 37°C under the standard assay method conditions.Refer to Table 1 for guidance on factors to adjust units according to temperature of assay.
Cholesterol Oxidase(Recombinant)
E N Z Y M E S
CAT# RECO-70-1229
EC# 1.1.3.6
C A T # R E C O - 7 0 - 1 2 2 9E C # 1 . 1 . 3 . 6
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ASSAY TEMPERATURE
25°C 0.75
30°C 0.86
37°C 1.00
45°C 1.02
50°C 1.01
FACTOR RELATIVE TO 37° RESULT
TABLE 1 : T EMPERATURE FACTORS FOR UN I T CONVERS ION
Note: Temperature can influence the level of available oxygen in the reaction mixture.
� A S S AY P R I N C I P L ECholesterol Oxidase (CO) catalyses the following reaction:
Cholesterol + O2 Cholestenone + H2O2CO
CAT# RECO-70-1229
EC# 1.1.3.6
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SUBS TRATE SPEC I F IC I TY
SUBSTRATE
Pregnenolone 104%
Cholesterol 100%
β-Cholestanol 81%
Stigmasterol 51%
Ergosterol 39%
% OF CHOLESTEROL ACTIVITY
SUBSTRATE
Cholesteryl Linoleate 0.02%
Cholecalciferol 0.001%
Androsterone 0.001%
Estradiol 0.0003%
% OF CHOLESTEROL ACTIVITY
Molecular Weight (by SDS Page): 58kD
Kmvalue (using Eadie-Hofstee): 5 x 10-6 M (Cholesterol)
Optimum pH (Fig. 1): pH 6.5 to 7.5
Optimum Temperature (Fig. 2): 45°C
Stable pH Range (Fig. 3): pH 4.5 to 9.0 (25°C for 20 hrs)
Thermal Stability (Fig. 4): Stable at 60°C and below (pH 7.0 for 15 mins)
Substrate specificity was tested in-house by replacing cholesterol with alternativesubstrates in the CO assay i.e. at 0.1mM concentration.
� C H A R A C T E R I S T I C SThis recombinant grade of CO is highly purified. Its main characteristics are as follows:
� A P P L I C A T I O NCholesterol Oxidase can be used in combination with Cholesterol Esterase (inbiosensors, test strips or other clinical chemistry formats) to determine the level ofcholesterol in blood. It is highly soluble over a wide pH.
F IGURE 3 : pH S TAB I L I TY
CAT# RECO-70-1229
EC# 1.1.3.6
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THE AMERICASSekisui Diagnostics, LLC4 Hartwell Place Lexington, MA 02421 Phone: 800 332 1042Fax: 800 762 6311
INTERNATIONALSekisui Diagnostics (UK) LimitedLiphook Way, AllingtonMaidstone, Kent, ME16 0LQ, UKPhone: +44 1622 607800Fax: +44 1622 607801
AP-00-0019.1.1 03/17© 2017 Sekisui Diagnostics, LLC. All rights reserved. Because every result matters™ is a trademark of Sekisui Diagnostics, LLC.
RELATIVE ACTIVITY (%)
pH
120
100
80
60
40
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0
3 4 5 6 7 8 9 10 11
F IGURE 1 : OP T IMUM pH
F IGURE 4 : T EMPERATURE S TAB I L I TY
F IGURE 2 : OP T IMUM T EMPERATURE
pH
3 4 5 6 7 8 9 10 11
RELATIVE ACTIVITY (%)
TEMPERATURE (°C)
TEMPERATURE (°C)
120
100
80
60
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25 30 35 40 45 50
RESIDUAL ACTIVITY (%)
120
100
80
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0
-20
RELATIVE ACTIVITY (%)
120
100
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30 35 40 45 50 55 60 65 70
Na ACETATE
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