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ANALIS
HS CEofix™ HbA2 kit for Beckman Coulter P/ACE™ 5000 series - GB: Instructions for use
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QUOTATION
INFORMATION
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| DESCRIPTION |
TECHNICAL DESCR. |
DESCRIPTION
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1. Date: 7-May-03-Revision: 01
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2. Intended use:
The kit is intended for the quantitative determination of human hemoglobin A2 (HbA2), determination of hemoglobin F and hemoglobin variants. For hemoglobin variants the kit should be used in combination with CEofix™ HbA1c kit.
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3. Principle of the test:
The hemoglobins are separated inside a capillary under the influence of an electrical field. Differences in charge to mass ratio give different migration times.
A UV detector is used and detects the hemoglobins at 415nm.
The kit consists of buffers and method. The buffers use the patented dynamic double coating technique.
The % HbA2 is defined as the percent of the HbA2 fraction in relation to the sum of all hemoglobin fractions.
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4. Content:
Rinse:1x100mL aqua bidistallata (PN 10-004183)
Conditioner:1x100mL NaOH 0.2 M (PN 10-004181)
Initiator:1x 2.5 mL Aqueous solution (PN 10-004194)
Buffer:4 x 4.2 mL Taurine Arginine pH 8.9 (PN 10-004192)
Hemolyser:1 x 5.2 mLArginine (PN 10-004195)
Micro vials:130
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5. Configuration of instrument:
-BeckmanCoulter P/ACE 5000
-UV detector with 415 nm filter (PN: 10-004709)
-P/ACE station software version 1.0.
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6. Other material needed:
- Vials (PN 358807)
- caps (PN 359079)
- spring (PN338488)
- Cartridge 100x800 µm ( PN 727610)
- Pipettes
- Capillary 25µM ID x 20cm to the detector, 31 cm total length (PN 10-004188/844111023)
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7. Training:
The operator should be familiar with instrument (P/ACE 5000) operation and maintenance.
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8. Reagent preparation and storage:
Reagents are ready to use. Store kit and buffers, in closed container, between 8°C and 30°C until expiration date.
Between operation buffer vials should be capped to avoid evaporation.
If the kit arrived damaged a replacement of the kit should be requested.
The kit contains no hazardous components.
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9. Type of specimen:
Biological fluids should be collected in the manner normally used for any clinical laboratory test.
Whole blood is the preferred specimen.
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11. Sample preparation:
Sample: 50 µL hemolyser + 10 µL of well-mixed whole blood in a microvial and mix.
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12. Operations:
Every day operation:
- clean instrument and interface manifold
- put new rinse, conditioner, initiator buffer vials- put new waste vial - put clean and dry caps on buffer vials
- run cond1.met when system was down for several days
- prepare samples
- place sample in instrument
- run HbA2.met method
End of day:
- remove samples
- cap buffer vials
Placing new capillary
-When the capillary is broken or when the capillary is not operating properly replace the capillary: according the instrument manual. -run cond1.met for conditioning
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13. Calibration:
No calibration is required for the kit.
Instrument should be maintained and calibrated according to Beckman Coulter specifications.
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15. Calculation:
% HbA2 = corrected area of HbA2 fraction in relation with the sum of corrected areas of all hemoglobin fractions.
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16. Imprecision:
Imprecision
| HbA2 |
HbA2 (2.9%) |
MT A2 (4.7 min) |
| Within-assay (N=20) %CV |
1.4 |
2.3 |
| Between-run (N=20 days) %CV |
1.8 |
2.9 |
Analis May 1998
Correlation
Fig. Comparison of Hb A2 quantification.
CZE and MAEC results (n 5 57); ( B), CZE and HPLC results (n 5 57)
Hb A2 CZE (%) = 1.233 Hb A2 HPLC - 0.2
Fig. Comparison of Hb F values obtained with HPLC and CZE.
Hb FCZE (%) = 1.118 Hb FHPLC + 0.4
F. Cotton et al.: Clinical Chemistry 45, No. 2, 1999
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17. Reference range:
Each laboratory should establish its own reference range to conform to patient population.
HbA2:
HbF:
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18. Interference:
- The presence of hemoglobin variants will limit the determination of HbA2.
-For determination of hemoglobin variants the kit should be used in combination with CEofix HbA1c kit.
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19. Quality Control:
Pooled control or commercially available control should be included in each run.
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20. References:
1 Cotton F, Lin C, Fontaine B, Gulbis B, Janssens J, Vertongen F. Evaluation of a Capillary
Electrophoresis Method for Routine Determination of Hemoglobins A2 and F. Clin Chem
1999;45:237-43
2 Lin C, Cotton F, Fontaine B, Gulbis B, Janssens J, Vertongen F. Capillary zone electrophoresis: an
additional technique for identification of hemoglobin variants. Hemoglobin 1999;23:97-109
3 Gerritsma J, Sinnige D, Drieze C, Sittrop B, Houtsma P, Hulshorst-Jansen N and Huisman W.
Quantitative and qualitative analysis of haemoglobin variants using capillary zone
electrophoresis. Ann Clin Biochem 2000;37:380-9
4 Mario N, Baudin B, Bruneel A, Janssens J, Vaubourdolle M. Capillary Zone Electrophoresis for
the Diagnosis of Congenital Hemoglobinopathies. Clin Chem 1999;45:285-8
Bibliography
1 Castagnola M, Messana I, Cassiano L, Rabino R, Rossetti D, Giardina B. The use of capillary
electrophoresis for the determination of hemoglobin variants. Electrophoresis 1995;16:1492-8
2 Hempe J, Craver R. Quantification of hemoglobin variants by capillary isoelectric focusing. Clin
Chem 1994;40:2288-95
3 Hempe J, Granger J, Craver R. Capillary isoelectric focusing of hemoglobin variants in the
pediatric clinical laboratory. Electrophoresis 1997;18:1785-95
4 Hempe J, Granger J, Warrier R, Craver R. Analysis of hemoglobin variants by capillary
isoelectric focusing. J Cap Elec 1997;4:131-5
5 Hempe J, Craver R. Separation of hemoglobin variants with similar charge by capillary
isoelectric focusing : Value of isoelectric point for identification of common and uncommon
hemoglobin variants. Electrophoresis 2000;21:743-48
6 Jenkins M, Hendy J, Smith I. Evaluation of hemoglobin A2 quantification assay and hemoglobin
variant screening by capillary electrophoresis. J Cap Elec 1997;004:137-43
7 Landers J. Clinical capillary electrophoresis. Clin Chem 1995;41:495-509
8 Lin C, Gulbis B, Delobbe E, Cotton F, Vertongen F. Separation of human globin chains by
micellar electrokinetic capillary chromatography. J Chromatogr B 1998;719:47-54
9 Lukens J, Lee R. Hereditary disorders affecting hemoglobin structure and synthesis. Wintrobe's
Clinical Hematology, Ninth Edition 1993;1:1023-145
10 Mario N, Baudin B, Aussel C, Giboudeau J. Capillary isoelectric focusing and high-performance
cation-exchange chromatography compared for qualitative and quantitative analysis of
hemoglobin variants. Clin Chem 1997;43:2137-42
11 Mohammad A et al. Clinical application of capillary isoelectric focusing on fused silica capillary
for determination of hemoglobin variants. Clin Chem 1997;43:1798-9
12 Molteni S, Frischknecht H, Thormann W. Application of dynamic capillary isoelectric focusing to
the analysis of human hemoglobin variants. Electrophoresis 1994;15:22-30
13 Oda R, Clark R, Katzmann J, Landers J. Capillary electrophoresis as a clinical tool for the
analysis of protein in serum and other body fluids. Electrophoresis 1997;18:1715-23
14 Ong C, Liau L, Ong H. Separation of globins using free zone capillary electrophoresis. J
Chromatogr 1992;576:346-50
15 Shihabi Z, Hinsdale M, Daugherty H. Hemoglobin A2 quantification by capillary zone
electrophoresis. Electrophoresis 2000;21:749-52
16 Sugano M et al. Analysis of hemoglobin and globin chain variants by a commonly used capillary
isoelectric focusing method. Electrophoresis 2000;21:3016-9.
17 Weykamp C, Penders T, Muskiet F, van der Slik W. Influence of hemoglobin variants and
derivatives on glycohemoglobin determinations, as investigated by 102 laboratories using 16
methods. Clin Chem 1993;39:1717-23
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TECHNICAL DESCR.
Revision 02
|
|
1. Date: 30-SEP-2004 -Revision: 02
|
|
2. Intended use:
The kit is intended for the quantitative determination of human hemoglobin A2 (HbA2), determination of hemoglobin F and hemoglobin variants. For hemoglobin variants the kit should be used in combination with CEofix™ HbA1c kit.
|
|
3. Principle of the test:
The hemoglobins are separated inside a capillary under the influence of an electrical field. Differences in charge to mass ratio give different migration times.
A UV detector is used and detects the hemoglobins at 415nm.
The kit consists of buffers and method. The buffers use the patented dynamic double coating technique.
The % HbA2 is defined as the percent of the HbA2 fraction in relation to the sum of all hemoglobin fractions.
|
|
|
4. Content:
Rinse:1x100mL aqua bidistallata (PN 10-004183)
Conditioner:1x100mL NaOH 0.2 M (PN 10-004181)
Initiator:1x 2.5 mL Aqueous solution (PN 10-004194)
Buffer:4 x 4.2 mL Taurine Arginine pH 8.7 (PN 10-004192)
Hemolyser:1 x 5.2 mLArginine (PN 10-004195)
Micro vials:130
|
|
|
5. Configuration of instrument:
-BeckmanCoulter P/ACE 5000
-UV detector with 415 nm filter (PN: 10-004709)
-P/ACE station software version 1.0.
|
|
|
6. Other material needed:
- Vials (PN 358807)
- caps (PN 359079)
- spring (PN338488)
- Cartridge 100x800 µm ( PN 727610)
- Pipettes
- Capillary 25µM ID x 20cm to the detector, 31 cm total length (PN 10-004188/844111023)
|
|
7. Training:
The operator should be familiar with instrument (P/ACE 5000) operation and maintenance.
|
|
|
8. Reagent preparation and storage:
Reagents are ready to use. Store kit and buffers, in closed container, between 8°C and 30°C until expiration date.
Between operation buffer vials should be capped to avoid evaporation.
If the kit arrived damaged a replacement of the kit should be requested.
The kit contains no hazardous components.
|
|
9. Type of specimen:
Biological fluids should be collected in the manner normally used for any clinical laboratory test.
Whole blood is the preferred specimen.
|
|
|
11. Sample preparation:
Sample: 50 µL hemolyser + 10 µL of well-mixed whole blood in a microvial and mix.
|
|
|
12. Operations:
Every day operation:
- clean instrument and interface manifold
- put new rinse, conditioner, initiator buffer vials- put new waste vial - put clean and dry caps on buffer vials
- run cond1.met when system was down for several days
- prepare samples
- place sample in instrument
- run HbA2.met method
End of day:
- remove samples
- cap buffer vials
Placing new capillary
-When the capillary is broken or when the capillary is not operating properly replace the capillary: according the instrument manual. -run cond1.met for conditioning
|
|
|
13. Calibration:
No calibration is required for the kit.
Instrument should be maintained and calibrated according to Beckman Coulter specifications.
|
|
|
15. Calculation:
% HbA2 = corrected area of HbA2 fraction in relation with the sum of corrected areas of all hemoglobin fractions.
|
|
|
16. Imprecision:
Imprecision
| HbA2 |
HbA2 (2.9%) |
MT A2 (4.7 min) |
| Within-assay (N=20) %CV |
1.4 |
2.3 |
| Between-run (N=20 days) %CV |
1.8 |
2.9 |
Analis May 1998
Correlation
Fig. Comparison of Hb A2 quantification.
CZE and MAEC results (n 5 57); ( B), CZE and HPLC results (n 5 57)
Hb A2 CZE (%) = 1.233 Hb A2 HPLC - 0.2
Fig. Comparison of Hb F values obtained with HPLC and CZE.
Hb FCZE (%) = 1.118 Hb FHPLC + 0.4
F. Cotton et al.: Clinical Chemistry 45, No. 2, 1999
|
|
17. Reference range:
Each laboratory should establish its own reference range to conform to patient population.
HbA2:
HbF:
|
|
|
18. Interference:
-.The presence of hemoglobin variants will limit the determination of HbA2.
-For determination of hemoglobin variants the kit should be used in combination with CEofix HbA1c kit.
|
|
|
19. Quality Control:
Pooled control or commercially available control should be included in each run.
|
|
|
|
20. References:
1 Cotton F, Lin C, Fontaine B, Gulbis B, Janssens J, Vertongen F. Evaluation of a Capillary
Electrophoresis Method for Routine Determination of Hemoglobins A2 and F. Clin Chem
1999;45:237-43
2 Lin C, Cotton F, Fontaine B, Gulbis B, Janssens J, Vertongen F. Capillary zone electrophoresis: an
additional technique for identification of hemoglobin variants. Hemoglobin 1999;23:97-109
3 Gerritsma J, Sinnige D, Drieze C, Sittrop B, Houtsma P, Hulshorst-Jansen N and Huisman W.
Quantitative and qualitative analysis of haemoglobin variants using capillary zone
electrophoresis. Ann Clin Biochem 2000;37:380-9
4 Mario N, Baudin B, Bruneel A, Janssens J, Vaubourdolle M. Capillary Zone Electrophoresis for
the Diagnosis of Congenital Hemoglobinopathies. Clin Chem 1999;45:285-8
Bibliography
1 Castagnola M, Messana I, Cassiano L, Rabino R, Rossetti D, Giardina B. The use of capillary
electrophoresis for the determination of hemoglobin variants. Electrophoresis 1995;16:1492-8
2 Hempe J, Craver R. Quantification of hemoglobin variants by capillary isoelectric focusing. Clin
Chem 1994;40:2288-95
3 Hempe J, Granger J, Craver R. Capillary isoelectric focusing of hemoglobin variants in the
pediatric clinical laboratory. Electrophoresis 1997;18:1785-95
4 Hempe J, Granger J, Warrier R, Craver R. Analysis of hemoglobin variants by capillary
isoelectric focusing. J Cap Elec 1997;4:131-5
5 Hempe J, Craver R. Separation of hemoglobin variants with similar charge by capillary
isoelectric focusing : Value of isoelectric point for identification of common and uncommon
hemoglobin variants. Electrophoresis 2000;21:743-48
6 Jenkins M, Hendy J, Smith I. Evaluation of hemoglobin A2 quantification assay and hemoglobin
variant screening by capillary electrophoresis. J Cap Elec 1997;004:137-43
7 Landers J. Clinical capillary electrophoresis. Clin Chem 1995;41:495-509
8 Lin C, Gulbis B, Delobbe E, Cotton F, Vertongen F. Separation of human globin chains by
micellar electrokinetic capillary chromatography. J Chromatogr B 1998;719:47-54
9 Lukens J, Lee R. Hereditary disorders affecting hemoglobin structure and synthesis. Wintrobe's
Clinical Hematology, Ninth Edition 1993;1:1023-145
10 Mario N, Baudin B, Aussel C, Giboudeau J. Capillary isoelectric focusing and high-performance
cation-exchange chromatography compared for qualitative and quantitative analysis of
hemoglobin variants. Clin Chem 1997;43:2137-42
11 Mohammad A et al. Clinical application of capillary isoelectric focusing on fused silica capillary
for determination of hemoglobin variants. Clin Chem 1997;43:1798-9
12 Molteni S, Frischknecht H, Thormann W. Application of dynamic capillary isoelectric focusing to
the analysis of human hemoglobin variants. Electrophoresis 1994;15:22-30
13 Oda R, Clark R, Katzmann J, Landers J. Capillary electrophoresis as a clinical tool for the
analysis of protein in serum and other body fluids. Electrophoresis 1997;18:1715-23
14 Ong C, Liau L, Ong H. Separation of globins using free zone capillary electrophoresis. J
Chromatogr 1992;576:346-50
15 Shihabi Z, Hinsdale M, Daugherty H. Hemoglobin A2 quantification by capillary zone
electrophoresis. Electrophoresis 2000;21:749-52
16 Sugano M et al. Analysis of hemoglobin and globin chain variants by a commonly used capillary
isoelectric focusing method. Electrophoresis 2000;21:3016-9.
17 Weykamp C, Penders T, Muskiet F, van der Slik W. Influence of hemoglobin variants and
derivatives on glycohemoglobin determinations, as investigated by 102 laboratories using 16
methods. Clin Chem 1993;39:1717-23
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Prices are quoted in euro (taxes excluded) and apply in the Benelux (recycling fee included). They are subject to change without notice. We reserve the right to adjust pricing errors and to limit quantities.
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