AEROSET® & ARCHITECT® cSystem™ MULTIGENT® Creatinine (Enzymatic) REF 8L24 Target Launch: August 2008

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

MULTIGENT® Creatinine (Enzymatic) Launch Strategy Two size codes of the reagent will be distributed from Chicago for use in the U.S. only. (Size codes 8L24-01 & 8L24-02 are distributed from Delkenheim to non-US countries, including Canada and Latin America.) Calibrator Clin Chem Calibrator ( REF 6K30-02) is available for use within the US. Controls Abbott will not market controls for U.S. use (Ranges for appropriate Bio-Rad controls have been established) Complaint handling: Abbott has responsibility for initial complaint registration. Sentinel (the vendor) will investigate level II complaints.

MULTIGENT® Creatinine (Enzymatic) Launch Strategy – Assay Disk MULTIGENT c Systems Assay Disk v2.0 will be available at assay launch REF 8K50-02 Conventional/Alternate Units REF 8K51-02 SI/Alternate units Creatinine (Enzymatic) reagent will contain a linearity addendum instructing the customer to change the Linear fields if the parameters have been loaded from the current ARCHITECT cSystems assay disk. No AEROSET disk will be produced. Linearity parameters for AEROSET users are correct in the insert. No edit is required.

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

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

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.

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 EDTA is not recommended. Internal testing showed decreased results - Mean % Bias of –9.1% at the medical decision level of 1.0 mg/dL. Urine Collection at <24 h with no preservatives Collection at 24 h with Boric Acid or HCl as preservatives Inspect for particulates

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

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

MULTIGENT® Creatinine (Enzymatic) Reagent Handling Kit Configuration REF 8L24-30 R1: 2 x 90 mL R2: 2 x 35 ml REF 8L24-40 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.

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

MULTIGENT® Creatinine (Enzymatic) Calibration Calibration stability is 60 days Uses MULTIGENT® Clin Chem Cal Calibrator kits currently contain enzymatic creatinine values. 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 6K30-02 4 x 3 mL

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

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

Quality Control Initial control ranges were communicated to Bio-Rad for the following products. Refer to the Bio-Rad insert for official labeling. Range for Crea Enz.was calculated using the following specs:  20% for serum controls (Rilibak German Directive for serum TAE = +/- 22%) ± 22% for urine controls (Rilibak German Directive for urine TAE = +/- 24%) TAE = total allowable error

MULTIGENT ® Creatinine (Enzymatic) Reportable Range Serum and Plasma: Reportable Range = 0.10 to 40.00 mg/dL (8.8 to 3,536.0 mol/L) Linear within  5% or 0.05 mg/dL whichever is greater Limit of Quantitation = 0.10 mg/dL (8.8 mol/L) Urine: Reportable Range = 2.50 to 400.00 mg/dL (0.22 to 35.36 mmol/L) Linear within 5% or 5 mg/dL whichever is greater Limit of Quantitation = 2.20 mg/dL (0.19 mmol/L)

MULTIGENT Creatinine (Enzymatic) Interference

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

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.

MULTIGENT® Creatinine (Enzymatic) Precision Urine precision: The precision of the Creatinine (Enzymatic) assay is < 3.6% Total CV for concentrations > 60.00 mg/dL and SD < 3.60 mg/dL for concentrations < 60.00 mg/dL. Representative data from a 20 day study based on Clinical and Laboratory Standards Institute (CLSI) protocol NCCLS EP5-A are summarized below.

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

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

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-11 0.69 0.697 CHM-12 5.27 5.173 CHM-13 7.30 7.105 CHM-14 3.92 3.889 CHM-15 1.34 1.36

ARCHITECT® Reaction graph (Low Range): CrEnz

ARCHITECT® Reaction graph (High Range): CrEnz

AEROSET® Reaction graph (Low Range): CrEnz

AEROSET® Reaction graph (High Range): CrEnz

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.

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.

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

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