Procedure:
Procainamide
OSR4K229
This procedure is valid for the following chemistry analyzers:
· AU400/AU400e / · AU640/AU640e· AU480 / · AU680
· AU600 / · AU2700
· AU5400 / · AU5800
Prepared By / Date Adopted / Supersedes Procedure #
Review Date / Revision Date / Signature
Distributed to / # of
Copies / Distributed to / # of
Copies
PRINCIPLE:
Procainamide is a cardiac anti-arrhythmic medication used to correct disturbances in the heart’s rhythm. Procainamide is used for therapy of PVCs, ventricular tachycardia, atrial fibrillation, and paroxysmal atrial tachycardia.
Anti-arrhythmic activity of procainamide correlates better with the drug's concentration in serum than with dosage. Therefore, monitoring serum procainamide concentration helps to achieve an optimum anti-arrhythmic effect and reduce the risk of toxicity.
Serum procainamide monitoring is particularly important because the range of procainamide serum concentrations required to achieve an effective and safe anti-arrhythmic response is narrow. In addition, patient-to-patient variations in absorption, distribution, metabolism, and elimination of procainamide make it difficult to predict serum levels from the administered dosage, particularly in patients with renal impairment.1,2
The presence of procainamide's pharmacologically active metabolite, N-acetyl- procainamide, complicates the definition of a range of effective serum procainamide concentrations. Since patients receiving procainamide for longer than 24 hours are in effect receiving two drugs, plasma N-acetylprocainamide levels should be measured along with procainamide levels.1,2
Methods historically used to monitor serum procainamide and N-acetyl- procainamide concentrations include immunoassay, gas-liquid chromatography (GLC) assay, and high-performance liquid chromatography (HPLC).1-3
INTENDED USE:
Emit® 2000 Procainamide Assay is intended for use in the quantitative analysis of procainamide in human serum or plasma. Emit® 2000 Assays are designed for use with multiple Beckman Coulter AU Analyzers.
METHODOLOGY:
The Emitâ 2000 Procainamide Assay is a homogeneous enzyme immunoassay technique used for the analysis of specific compounds in biological fluids.4,5 The assay is based on competition between drug in the sample and drug labeled with the enzyme glucose-6-phosphate dehydrogenase (G6PDH) for antibody binding sites. Enzyme activity decreases upon binding to the antibody, so the drug concentration in the sample can be measured in terms of enzyme activity. Active enzyme converts oxidized nicotinamide adenine dinucleotide (NAD) to NADH, resulting in an absorbance change that is measured spectrophotometrically. Endogenous serum G6P-DH does not interfere because the coenzyme functions only with the bacterial (Leuconostoc mesenteroides) enzyme employed in the assay.
SPECIMEN:
Patient / Sample Preparation:
No special preparation for the patient is required. The patient’s clinical condition and dosage regimen may influence the sample collection time.
Pharmacokinetic factors influence the correct time of sample collection after the last drug dose. These factors include dosage form, mode of administration, concomitant drug therapy, and biological variations affecting drug disposition.1,2
Sample Collection Time:
Measure the steady-state serum concentration representing the trough level just before the next scheduled dose.
Additional instructions for preparation as designated by this laboratory:Type:
Serum or plasma is the recommended specimen. Whole blood cannot be used. The anticoagulants heparin, citrate, oxalate, and EDTA have been tested and may be used with this assay.
Some sample dilution may occur when samples are collected in tubes containing citrate anticoagulant. The amount of dilution and the possible need to correct for it should be considered when interpreting assay results for these samples.
Additional type conditions as designated by this laboratory:Handling Conditions:
Store the serum or plasma refrigerated at 2-8°C. For transporting, maintain the sample temperature at 2-8°C. Samples can be stored refrigerated at 2-8°C for up to 7 days or stored frozen (-20°C) for up to one month.
Samples that contain particulate matter, fibrous material, or gel-like masses; appear unusual; or are frozen require preparation. Use the following instructions to prepare such samples:
1. If sample is frozen, thaw at a room temperature of 15-25°C.
2. Vigorously mix all samples: Vortex for at least 30 seconds.
3. Centrifuge sample at 2000 rpm for 15 minutes.
4. Collect a sample from the middle portion of the specimen. Avoid collecting lipids from the top portion or particulate matter from the bottom portion.
Human serum or plasma samples should be handled and disposed of as if they were potentially infectious.
Additional handling conditions as designated by this laboratory:EQUIPMENT AND MATERIALS:
Equipment:
Beckman Coulter AU400/AU400e, AU480, AU600, AU640/AU640e, AU680, AU2700, AU5400 AU5800 analyzers.
Materials:
Emitâ 2000 Procainamide Assay
Antibody/Substrate Reagent 1 — mouse monoclonal antibodies reactive to procainamide, glucose-6-phosphate, nicotinamide adenine dinucleotide, preservatives, and stabilizers.
Enzyme Reagent 2 — procainamide labeled with glucose-6-phosphate dehydrogenase, Tris buffer, preservatives, and stabilizers.
Reagent storage location in this laboratory:Test tubes 12 -16 mm in diameter or sample cups (Cat No. AU1063)
Storage location of test tubes or sample cups in this laboratory:Emitâ 2000 Procainamide Calibrators (Cat No. 4K109)
The Emitâ 2000 Procainamide Calibrators contain the following stated procainamide concentrations: 0 mg/mL, 1.0 mg/mL, 2.0 mg/mL, 4.0 mg/mL, 8.0 mg/mL, and 16 mg/mL. The calibrator kit is sold separately.
Storage location of the calibrator in this laboratory:Preparation
The Emitâ 2000 Procainamide reagents and calibrators are packaged in a ready to use liquid form and may be used directly from the refrigerator. Close the calibrator vials when not in use. Caps must always be replaced on the original containers.
Note: Reagents 1 and 2 are provided as a matched set. They should not be interchanged with components of kits with different lot numbers.
Precautions:
1. The Emit® 2000 Procainamide reagents and calibrators are for in vitro diagnostic use.
2. Reagent 1 contains non-sterile mouse monoclonal antibodies.
3. Do not use the reagents or calibrators after the expiration date.
Storage Requirements:
Any reagents not loaded in the reagent refrigerator on the analyzer or any calibrators not in use should be store at 2-8°C (36-46°F), upright, and with caps tightly closed. Do not freeze reagents or calibrators or expose them to temperatures above 32°C.
Unopened reagents and calibrators are stable until the expiration date printed on the label if stored as directed. Refer to Assay Methodology Sheets for additional on-board stability information.
Improper storage of reagents or calibrators can affect assay performance. Stability depends on handling reagents or calibrators as directed.
Additional storage requirements as designated by this laboratory:Indications of Deterioration:
Discoloration (especially yellowing) of the reagents or calibrators, visible signs of microbial growth, turbidity, or precipitation in reagent or calibrators may indicate degradation and warrant discontinuance of use.
PERFORMANCE PARAMETERS:
The following performance characteristics represent total system performance and should not be interpreted to refer only to reagents. Studies were performed on the Beckman Coulter AU analyzer series. Results may vary due to analyzer-to-analyzer differences.
Precision
Within run precision was calculated by running twenty replicates of each level of a tri-level control. Total precision was calculated according to Clinical and Laboratory Standards Institute (CLSI EP5-A) using data collected from controls run in duplicate twice daily over twenty days (N=80).
Within-Run Precision / Total PrecisionLevel 1 / Level 2 / Level 3 / Level 1 / Level 2 / Level 3
Mean (µg/mL) / 2.47 / 6.91 / 12.25 / 2.41 / 6.74 / 11.13
CV % / 1.3 / 1.7 / 4.8 / 3.7 / 3.6 / 7.9
Comparison
Samples from patients were analyzed using the Emit® 2000 Procainamide Assay on the TDx analyzer and the AU600. Results are shown below.
Slope / 1.00Intercept (mg/mL) / 0.02
Mean (mg/mL)
TDx
AU600 / 4.67
4.71
Correlation Coefficient / 0.995
Number of Samples / 54
CALIBRATION:
Perform a multi-point calibration (5AB) using a water blank (blue rack) and the Emit® 2000 Procainamide Calibrators: 1.0, 2.0, 4.0, 8.0, 16.0. Calibration parameters are set to prepare the calibration curve. Refer to analyzer User’s Guide or Analyzer Specific Protocol sheets for analyzer settings.
Calibration Stability
Studies have shown the median calibration stability to be at least 14 days. Recalibrate as indicated by control results or with a new lot of reagent. Calibration stability may vary from laboratory to laboratory depending on the following: handling of reagents, maintenance of analyzer, adherence to operating procedures, establishment of control limits, and verification of calibration.
Note: When using a new set of reagents with the same lot number, validate the system by assaying controls.
QUALITY CONTROL:
During operation of the Beckman Coulter AU analyzer multi-level controls should be tested in every run or a minimum of once a day. Controls should be performed after calibration, with each new set or lot of reagent, and after specific maintenance or troubleshooting steps described in the appropriate User’s Guide. Quality control testing should be performed in accordance with regulatory requirements and individual laboratory’s standard procedures. If more frequent verification of test results is required by the operating procedures within your laboratory, those requirements should be met.
PARAMETERS
A complete list of test parameters and operating procedures can be found in the appropriate User’s Guide and at www.beckmancoulter.com.
CALCULATIONS:
Results are calculated automatically by the analyzer. No additional manipulation of data is required.
This assay uses Math Model No. 1.
To convert from mg/mL to mmol/L procainamide, multiply by 4.25.
REPORTING RESULTS:
Reference Ranges:
The desired therapeutic effect is usually achieved in the serum concentration range of 4.0-10 mg/mL (17-43 mmol/L) and the sum of procainamide and N-acetylprocainamide concentrations is 10-30 mg/mL.1,2 Further, peak concentrations of procainamide above 12 mg/mL (51 mmol/L) are often associated with toxicity.1
For effective treatment, some patients may require serum levels outside this range. Therefore, the expected range is provided only as a guide, and individual patient results should be interpreted in light of other clinical signs and symptoms.
Expected reference ranges in this laboratory:Procedures for Abnormal Results
The laboratory must define procedures to be used in reporting high concentration (toxic) results to the patient’s physician.
Abnormal results are flagged by the listed analyzers according to the normal values entered by the user into the analyzer parameters
Reporting Format:
Results are automatically printed for each sample in mg/mL at 37°C.
Interpretation of Results
The factors that can influence the relationship between the procainamide serum or plasma concentrations and clinical response include renal and circulatory function, rate of acetylation, the type and severity of cardiac arrhythmia, general state of health, and use of other drugs.
The concentration of procainamide in serum or plasma depends on the time of the last drug dose; mode of administration; concomitant drug therapy; sample condition; time of sample collection; and individual variations in absorption, distribution, biotransformation, and excretion. These parameters must be considered when interpreting results.1 2
Additional reporting information as designated by this laboratory:LIMITATIONS:
The Emit® 2000 Procainamide Assay accurately quantitates procainamide concentrations in human serum or plasma containing 1.0-12 mg/mL (4.3-51 mmol/L) procainamide.
To estimate procainamide concentrations above the assay range, patient samples containing more than 12 mg/mL (51 mmol/L) procainamide may be diluted with one or two parts distilled or deionized water or Emitâ 2000 Procainamide Calibrator 0. After diluting the sample, repeat the entire assay sequence and multiply the results by the dilution factor.
Adulteration of reagents, use of analyzer without appropriate capabilities, or other failures to follow instructions as set forth in this protocol or the package insert can affect performance characteristics and stated or implied claims.
Interfering Substances
No clinically significant interference has been found in samples to which 800 mg/dL hemoglobin, 1000 mg/dL triglycerides, or 30 mg/dL bilirubin were added to simulate hemolytic, lipemic or icteric samples.
Procaine cross-reacts with the assay.
Sensitivity
The sensitivity level of the Emit® 2000 Procainamide Assay is 0.25 mg/mL. This level represents the lowest measurable concentration of procainamide that can be distinguished from 0 mg/mL with a confidence level of 95%.
Specificity
The Emitâ 2000 Procainamide Assay measures the total (protein-bound plus unbound) procainamide concentration in serum or plasma. Compounds whose chemical structure or concurrent therapeutic use would suggest possible cross-reactivity have been tested. Of the compounds tested, only procaine cross-reacts with the assay.
The compounds listed in the following table do not interfere with the Emitâ 2000 Procainamide Assay when tested in the presence of 4.0 mg/mL procainamide. Levels tested were at or above maximum physiological or pharmacological concentrations.
Compound / Concentration Tested(mg/mL)
Acetaminophen / 100
Desethyl-n-acetylprocainamide (DENAPA) / 100
Digoxin / 0.1
Diphenylhydantoin / 100
Disopyramide / 100
Ephedrine / 100
Furosemide / 100
Glycinexylidide (GX) / 100
Hydrochlorothiazide / 100
Isoproterenol / 100
Lidocaine / 100
Monoethylglycinexylidide (MEGX) / 100
N-Acetylprocainamide / 40
N-(2-Diethylaminoethyl) isonicotinamide / 100
p-Acetamidobenzoic acid / 100
p-Aminobenzoic acid / 100
Propranolol / 100
Quinidine / 100
Tocainide / 100
REFERENCES:
1. Lima JJ, Lewis RP. Procainamide: Therapeutic Use and Serum Concentration Monitoring. In: Taylor WJ, Finn AL eds. Individualizing Drug Therapy: Practical Applications of Drug Monitoring. New York: Gross, Townsend, Frank, Inc. 1981, vol 3, pp 69-88.
2. Coyle JD, Lima JJ. Procainamide. In: Evans WE, Schentag JJ, Jusko WJ eds. Applied Pharmacokinetics: Principles of Therapeutic Drug Monitoring. Washington: Applied Therapeutics, Inc. 1986, pp 682-711.
3. Bauer LA, Black D, Gensler A, et al. Influence of age, renal function, and heart failure on procainamide clearance and n-acetylprocainamide serum concentrations. Int J Clin Pharmacol Ther Tox. 1989; 27(5):213-216.
4. Pincus MR, Abraham NZ Jr. Toxicology and Therapeutic Drug Monitoring. In: Henry JB ed. Clinical Diagnosis and Management by Laboratory Methods, Ed 18. Philadelphia: WB Saunders Co. 1991, pp 349–384.
5. Lam X, Hefner A: Syvaâ Emit® 2000 Procainamide Assay. Clin Chem. 1992; 38(6):337. Abstract.
© Beckman Coulter, Inc. March 2012 CLSIOSR4K229.02
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