Albumin comprises about 60 percent of the total serum protein in a normal, healthy individual. It serves a number of functions, including transport of large insoluble organic anions (e.g. long chain fatty acids and bilirubin), binding of toxic heavy metal ions, transport of excess quantities of poorly soluble hormones (e.g. cortisol, aldosterone and thyroxine), maintenance of serum osmotic pressure, and provision of a reserve store of protein. The binding ability of albumin is the basis of many of the quantitative assays developed since the 1950s. This method utilizes the dye bromcresol purple (BCP). When the dye binds selectively with albumin in a pH range of 5.2-6.8, a color change occurs that is measured at 600 nm. The secondary wavelength is 700 nm. This is a 2-point, endpoint reaction that is specific for albumin. Cobas 6000 Application Code: 760
SPECIMEN COLLECTION, STORAGE, AND HANDLING PROCEDURES; CRITERIA FOR SPECIMEN REJECTION
a. Specimen Type and Requirements: Serum, and EDTA- or heparin-anticoagulated plasma are acceptable for this procedure. The NHANES Biochem study uses refrigerated serum. This test is analyzed from NHANES Vial 018.
b. Specimen Volume: Optimum/Minimum volume: 100 μL in a sample cup or 2 mL microtube (2 μL for test; remainder for dead volume).
c. Acceptable Specimens/Unacceptable Specimens: Serum, and EDTA- or heparinanticoagulated plasma. A fasting specimen is desirable. Other anticoagulants are not acceptable.
d. Specimen Stability and Storage: Separated serum or plasma should be removed from the cells within one hour of collection. Separated serum or plasma is stable for five days at 4ºC, four months at -20°C, and longer at –70ºC
e. Interferences or limitations: Icteric index <60: no interference. Hemolytic index <1000: no interference. Lipemic index <1500: no interference. Drugs (therapeutic concentrations of commonly used drug panels): no interference. In very rare cases, gammopathy, in particular IgM (Waldenstrom’s macroglobulinemia), may cause unreliable results. Rheumatoid factors <1200: no interference
f. Specimen Handling and Transport: If possible, the patient should be in a standardized body position (e.g., sitting) for at least 15 minutes prior to specimen collection. Erect posture causes a redistribution of body fluids resulting in increased serum albumin concentration and recumbent tends to lower the albumin concentration. Mix specimens well, allow clot to fully form (if serum), and centrifuge 10 minutes at 2000 x g before use. Aliquot a minimum of 0.1 mL. Store sample in refrigerator until shipment. Ship at refrigerated temperature.
EQUIPMENT AND INSTRUMENTATION, MATERIALS, REAGENT PREPARATION, CALIBRATORS (STANDARDS), AND CONTROLS
a. Reagents and Supplies
• Roche product #05599261190, ALBP reagent kit (225 tests):
b. Reagent Preparation (*Reagents are ready to use; no preparation required)
• R1 reagent. Buffer, preservatives, surfactants. No preparation required.
• R2 reagent. Buffer, preservatives, surfactant, bromcresol purple. See insert for concentrations. No preparation required.
Storage and stability. Keep reagents stored at room temperature until use. The reagents are stable for 28 days refrigerated on the analyzer.
c. Equipment/Instrumentation-
• Roche Cobas 6000 Chemistry Analyzer (Roche Diagnostics Corporation, Indianapolis, IN 46250)
• The Millipore Elix Gulfstream Clinical 35 System is designed to meet CLSI Clinical Laboratory Reagent Water (CLRW) standards. Water purification is achieved by reverse osmosis, electrodeionization, bactericidal 254 nm UV lamp and 0.22 μm filtration.
d. Specimens are run in singleton
e. Quality Control
• Normal pooled serum control (CQ). Stable at -80°C for up to 4 years, at refrigerated temperature for up to 1 day and at room temperature for up to 4 hours.
• Roche Precipath U Plus Control (catalog #12149443160). Roche Diagnostics 9115 Hague Road Indianapolis, IN 46250-0457). Stable until expiration date on package when unopened and stored at 2-8°C. To prepare, open bottle 1 and pipette in exactly 3.0 mL of diluent (bottle 2). Dissolve by gentle swirling for 30 minutes. Prepared control is stable for 12 hours at room temperature, 5 days at 2-8°C, and one month at -20°C (when frozen once).
Reference Range: Serum or plasma, adult: 3.5-5.0 g/dL
Limit of Detection (standard 1 + 3 SD): 0.3 g/dL
. SPECIMEN STORAGE AND HANDLING DURING TESTING
Specimens are stored at refrigerated temperature between sample receipt and analysis on the instrument. Specimens must be at room temperature prior to assay. Specimens are returned to refrigerated or frozen temperature post analysis depending on the study specific requirements.
LIMITATIONS OF METHOD; INTERFERING SUBSTANCES AND CONDITIONS
a. Limit of Detection (standard 1 + 3 SD): 0.3 g/dL
Analytical Measurement Range: 0.3-10 g/dL
b. Icteric index <60: no interference. Hemolytic index <1000: no interference. Lipemic index <1500: no interference. Drugs (therapeutic concentrations of commonly used drug panels): no interference. In very rare cases, gammopathy, in particular IgM (Waldenstrom’s macroglobulinemia), may cause unreliable results. Rheumatoid factors <1200: no interference
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