1. AEROSET® & ARCHITECT® cSystem™
MULTIGENT® Creatinine (Enzymatic) REF 8L24
Target Launch: EU April 2008

2. Agenda Launch Strategy Clinical Utility Methodology Sample Handling
Reagent Handling
Reagent SmartWash
Calibration
Quality Control
Reportable Range
Interference
Precision
Method Comparison
CAP Performance
Reaction Graphs
Troubleshooting tips
Questions and Answers

3. MULTIGENT® Creatinine (Enzymatic) Launch Strategy
Two size codes of the reagent will be distributed through Delkenheim to countries outside the U.S.
Calibrator
Clin Chem Calibrator ( REF 6K3010) is available for E.U. use.
Controls
Clin Chem Control 1 ( REF 6K3020) and Clin Chem Control 2 ( REF 6K3021) are available for E.U. use.
Complaint handling:
Abbott has responsibility for initial complaint registration.
Sentinel (the vendor) will investigate level II complaints.

4. MULTIGENT® Creatinine (Enzymatic) Launch Strategy – Assay Disk
A disk will be available at assay launch
The disk will be available in conventional and SI units versions.
No AEROSET disk will be produced.

5. Clinical Utility of Creatinine
Creatinine is eliminated from blood by glomerular filtration. Reduced renal function results in an increased serum creatinine concentration.
Measurement of serum creatinine is used to diagnose and monitor acute and chronic renal disease, estimate glomerular filtration rate (GFR), or assess the status of renal dialysis patients.
Urine creatinine analysis is used to calculate creatinine clearance, confirm completeness of 24-hour collections, or serve as a reference quantity for other analytes, such as in calculation of the albumin/ creatinine ratio

6. MULTIGENT® Creatinine (Enzymatic) Methodology
Methodology: Enzymatic
Chemical Reaction:
Creatinine + H2O Creatine
Creatine + H2O Sarcosine + Urea
Sarcosine + O2 + H2O Glycine Formaldehyde + H2O2
H2O2 + 4-AAP* + ESPMT** Quinoneimine Dye + 4H2O
* AAP = 4-aminoantipyrine
** ESPMT = N-ethyl-N-sulfopropyl-m-toluidine
Creatininase
Creatinase
Sarcosine Oxidase
Peroxidase

7. MULTIGENT® Creatinine (Enzymatic) Methodology
Description of the Enzymatic Reaction:
Creatinine in the sample is hydrolyzed by creatininase to creatine.
Creatine is in turn hydrolyzed by creatinase to sarcosine and urea.
Sarcosine from this reaction is oxidized by sarcosine oxidase to glycine and formaldehyde with the concomitant production of hydrogen peroxide
The hydrogen peroxide reacts with 4-aminoantipyrine and ESPMT* in the presence of peroxidase to yield a quinoneimine dye. The resulting change in absorbance at 548 nm is proportional to the creatinine concentration in the sample.
This enzymatic method is sensitive and specific for creatinine and is not affected by endogenous substances, such as ketoacids, cephalosporins, and bilirubin that interfere with the Jaffe method.

8. MULTIGENT® Creatinine (Enzymatic) Sample Handling
Specimen Requirements:
Serum
Plastic tubes
With or without gel barrier
Glass tubes were not tested
Plasma
Lithium Heparin, with or without barrier
Sodium Heparin
Do NOT use EDTA. Internal testing showed decreased results.
Urine
Collection at <24 h with no preservatives
Collection at 24 h with Boric Acid or HCl as preservatives
Inspect for particulates

9. MULTIGENT® Creatinine (Enzymatic) Sample Handling
Sample Tubes tested and found acceptable:
Vendor
REF
Tube
BD Europe
369032
Serum tube
367869
Na - Heparin
368884
Li – heparin (17 IU/mL)
367957
Serum with Gel separator
367374
Li-heparin with PST II gel barrier

10. MULTIGENT® Creatinine (Enzymatic) Sample Handling
Specimen storage:
Stored specimens must be inspected for particulates. If present, mix and centrifuge the specimen to remove particulates prior to testing.
Serum Sample volume = 4 uL
Urine Sample volume = 20 uL
System performs a Standard sample dilution of 1:10 using 20 uL of sample and 180 uL of saline diluent
Maximum Storage
Temperature
Serum/Plasma
Urine
20 to 25 C
7 days
2 days
2 to 8 C
6 days
-20 C
3 months
6 months

11. MULTIGENT® Creatinine (Enzymatic) Reagent Handling
Kit Configuration
REF 8L24-01
R1: 2 x 90 mL
R2: 2 x 35 ml
REF 8L24-02
R1: 9 x 90 mL
R2: 9 x 35 ml
Reagent is liquid, ready-to-use, two-reagent kit.
Storage: 2 to 8°C.
Unopened kit is stable until the expiration date.
Remove any air bubbles with an applicator stick.
Reagent onboard stability is 60 days.

12. Reagent SmartWash
The R1 reagent for Creatinine (Enzymatic) contains gentamicin. The following wash is required for the Gentamicin assay on ARCHITECT c Systems only.

13. MULTIGENT® Creatinine (Enzymatic) Calibration
Calibration stability is 60 days
Uses MULTIGENT® Clin Chem Cal
Calibrator kits currently contain enzymatic
creatinine values.
Creatinine (Enzymatic) reagent kit stuffer will contain Creatinine value assignments for existing lots of Clin Chem. Calibrator and Clin. Chem. Controls 1 and 2.
Creatinine (Enzymatic) is standardized to NIST standard reference material (SRM) 967. Values for SRM 967 are determined by isotope dilution mass spectrometry (IDMS).
Kit Configuration
REF 6K3010
4 x 3 mL

14. MULTIGENT ® Creatinine (Enzymatic) Calibration Curve - ARCHITECT®
CrEnz

15. MULTIGENT ® Creatinine (Enzymatic) Calibration Curve -AEROSET®
CrEnz

16. Quality Control
The following controls have been tested and initial control ranges have been assigned for AEROSET and ARCHITECT
Bio-Rad Liquid Assayed Multiqual Controls,
Lot # 45540, Exp. Date: 01/31/2009
Lot # 45550, Exp. Date: 09/30/2009
Bio-Rad Lyphochek Assayed Chemistry Controls,
Lot # 14140, Exp. Date: 12/31/2008
Lot # 14150, Exp. Date: 09/30/2009
Bio-Rad Liquichek Urine Chemistry Controls,
Lot # 62810, Exp. Date: 09/30/2008
Lot # 62820, Exp. Date: 02/28/2009
Lot # 62830, Exp. Date: 05/31/2009

17. Quality Control REF Configuration Clin Chem Control 1 6K3020 6 x 5 mL
The Sentinel manufactured ClinChem Controls are available for use.
REF
Configuration
Clin Chem Control 1
6K3020
6 x 5 mL
Clin Chem Control 2
6K3021

18. MULTIGENT ® Creatinine (Enzymatic) Reportable Range
Serum and Plasma:
Reportable Range = 0.14 to mg/dL (12.4 to 4,420.0 mol/L)
Linear within  5% or 0.05 mg/dL whichever is greater
Limit of Quantitation = 0.05 mg/dL (4.4 mol/L)
Urine:
Reportable Range = 3.00 to mg/dL (0.27 to mmol/L)
Linear within 5% or 5 mg/dL whichever is greater
Limit of Quantitation = 3.00 mg/dL (0.27 mmol/L)

19. MULTIGENT Creatinine (Enzymatic) Interference

20. MULTIGENT Creatinine (Enzymatic) Interference
Alpha-methyldopa is the only drug in the Creatinine (Enzymatic) serum application that showed interference >8%

21. MULTIGENT® Creatinine (Enzymatic) Precision
Serum precision:
The precision of the Creatinine (Enzymatic) assay is < 3.6% Total CV for concentrations >1.00 mg/dL and SD < 0.05 mg/dL for concentrations < 1.00 mg/dL.
Representative data from a 20 day study based on Clinical and Laboratory Standards Institute (CLSI) protocol NCCLS EP5-A are summarized below.

22. MULTIGENT® Creatinine (Enzymatic) Precision
Urine precision:
The precision of the Creatinine (Enzymatic) assay is < 3.6% Total CV for concentrations > mg/dL and SD < 3.60 mg/dL for concentrations
< mg/dL.
Representative data from a 20 day study based on Clinical and Laboratory Standards Institute (CLSI) protocol NCCLS EP5-A are summarized below.

23. MULTIGENT® Creatinine (Enzymatic) Method Comparison
SERUM
Results from the MULTIGENT Creatinine (Enzymatic) assay on the ARCHITECT c 8000 System and the AEROSET System were compared with the results from Sentinel Enzymatic Creatinine Assay on Hitachi 911.
Results from the MULTIGENT Creatinine (Enzymatic) assay on the ARCHITECT c 8000 System were compared with the results from REF 7D64 Creatinine on the ARCHITECT c 8000 system

24. MULTIGENT® Creatinine (Enzymatic) Method Comparison
URINE
Results from the MULTIGENT Creatinine (Enzymatic) assay on the ARCHITECT c 8000 System and the AEROSET System were compared with the results from Sentinel Enzymatic Creatinine Assay on Hitachi 911.
Results from the MULTIGENT Creatinine (Enzymatic) assay on the ARCHITECT c 8000 System were compared with the results from REF 7D64 Creatinine on the ARCHITECT c 8000 system

25. Creatinine (Enzymatic) CAP Performance
CAP Proficiency Testing (PT) for MULTIGENT Creatinine (Enzymatic) assay was performed in-house on the Architect c8000 analyzer using CAP PT materials : CHM-11, CHM-12, CHM-13, CHM-14 and CHM-15.
2007 CZ-C CAP Survey Results (Information Only) MULTIGENT All Method Mean
CHM
CHM CHM
CHM
CHM

26. ARCHITECT® Reaction graph (Low Range):
CrEnz

27. ARCHITECT® Reaction graph (High Range):
CrEnz

28. AEROSET® Reaction graph (Low Range):
CrEnz

29. AEROSET® Reaction graph (High Range):
CrEnz

30. Troubleshooting Tips
The R2 reagent for Creatinine (Enzymatic) is provided in Sentinel’s 50 mL trapezoid cartridge. Ensure that the cartridge is seated tightly and vertically in the reagent carousel to prevent potential for probe crashes or liquid level sense errors.

31. Questions and Answers
How does Creatinine measurement by Enzymatic method compare to Jaffe’s alkaline picrate method?
Enzymatic Creatinine measurement is sensitive and specific for creatinine and is not affected by endogenous substances, such as ketoacids, cephalosporins, and bilirubin that interfere with the Jaffe method.

32. Questions and Answers
The package insert indicates boric acid and hydrochloric acid are preferred preservatives. Are these the only ones that can be used?
No, Boric Acid and Hydrochloric Acid are the preferred preservatives. In addition, the other preservatives tested and found acceptable for use are:
Acetic Acid (8.5N) to 10 mL/dL
Nitric Acid (6N) to 6 mL/dL
Sodium Carbonate to 1.5 g/dL
Sodium Oxalate to 70 mg/dL
Sodium Fluoride to 500 mg/dL

33. Questions and Answers Are pediatric reference ranges available?
Refer to the troubleshooting guides which list ranges published by Ceriotti et al.1 based on the data of Schlebusch et al.2 or the original articles.
1 Ceriotti F, Boyd JC, Klein G, et al. Reference intervals for serum creatinine concentrations: assessment of available data for global application. Clin Chem 2008;54(3):559–66.
 2 Schlebusch H, Liappis N, Kalina E, et al. High sensitive CRP and creatinine: reference intervals from infancy to childhood. J Lab Med 2002;26(5/6):341–6.

34. Questions and Answers Was any commercial linearity material evaluated?
The following material was tested and found to be acceptable.