Total Cholesterol Examination Procedure
1.Purpose of Examination: To lay down standard operating procedure for serum Total Cholesterol estimation by CHOD-PAP Method in Biochemistry section of LSSTH, Bhavnagar.
2.Principle of the procedure used for examinations: The Cholesterol2 assay is an automated clinical chemistry assay. Cholesterol esters are enzymatically hydrolyzed by cholesterol esterase to cholesterol and free fatty acids. Free cholesterol, including that originally present, is then oxidized by cholesterol oxidase to cholest-4-ene-3-one and hydrogen peroxide. The hydrogen peroxide oxidatively couples with N,N-Bis(4-sulfobutyl)-3-methylaniline (TODB) and 4-aminoantipyrine to form a chromophore (quinoneimine dye) which is quantitated at 604 nm.
3.Performance: Linearity: upto 705mg/dl Limit of Detection (LOD):Serum : 5.0 mg/dl Limit of Quantitation (LOQ),:Serum :6.2 mg/dl The LOQ is the analyte concentration at which the CV = 20%.
4.Type of Primary Sample: Serum or Plasma sample_collection_manual
5.Patient Preparation: Instruction should be given to the patient Verbal consent of patient should be taken before collecting blood sample. Patient should relax for about 5 minutes before the venepuncture..
6.Type of Container and additives : Plain Vaccutainer with no additives
7.Required Equipment and reagents: Centrifuge, Auto-Pipette, Disposable Tips, Disposable sample cups, Fully Auto Chemistry Analyzer
8.Enviromental & Safety control: Reagent R1 contains sodium azide. contact with acids liberates very toxic gas
9.Precautions: Wear protective gloves / protective clothing / eye protection Do not breathe mist / vapors / spray Wash hands thoroughly after handling Keep only in original container
10.Calibration Procedure: Give Calibration with Consolidated Chemistry Calibrator ConCC (See calibrator chart for lot specific concentration) Manual calibration is done on following circumstances:
Note: Verify calibration with at least two levels of controls according to the established quality control requirements for your laboratory. If control results fall outside acceptable ranges, recalibration may be necessary
11.Procedural Steps:
12.Quality Control Procedure: The Laboratory runs Lypocheck Assayed Chemistry Control level 1 & level 2 Once a day. Monthly L-J charts are reviewed & for any QC outlier root cause analysis are done & corrective actions are taken according to the error. The Laboratory Participates in EQAS programme EQAS report is reviewed and if any result go outlier, then root cause analysis is done & corrective actions are taken according to the error.
13.Interferences & Cross-Reactions: InterferingSubstance
14.Principle of Procedure for Calculating Results including where relevant, the measurement uncertainty of measured quantity values: Instrument measures the absorbance of a calibrator of known concentration. This point determines the slope of a straight line that relates absorbance to concentration. Instrument calculates a constant which is the inverse of the slope of the straight line. Sample Concentration: Cs = K x As Cs = Sample Concentration K = Constant As = Sample Absorbance Measurement uncertainty: Highest CV% of last 6 months ± 1.96 %CV Refer Annexure I
15.Biological Reference Interval: Adult : <200mg/dl.
16.Reportable interval of patient examination results: Up to 500mg/dl
17.Manual Dilution Procedure:
18.Stability of sample Serum or plasma-At Room temperature18°–28°C (64°–82°F) stability ≤ 24 hours
19.Storage of sample 2-8° C for 1 days (Retention period for re-examination and/or additional tests is 24 hrs.)
20.Reagent Storage and stability Reagent is stored at 2-8 °c.Reagent stability is 30 days if the reagent is uncapped and on board.
21.Turn around time (TAT) 6.0 hours after collection,in case of emergency 2.0 hours
22.Critical Value:NA
23.Laboratory Interpretation: The common causes of high cholesterol level are as follows :hyperlipoproteinemias,stress,hormonal imbalance etc
24.Potential source of variation:
25.Reagent Handling Remove any air bubbles present in the reagents with a new applicator stick, or allow the reagents to sit at the appropriate storage temperature to allow the bubbles to dissipate. To minimize volume depletion, do not use a transfer pipette to remove bubbles
26.REFERENCE:
| Printed copy of this document is considered uncontrolled. It should be compared with controlled electronic copy before use |
| Name of Laboratory : Laboratory Services Sir T. Hospital (LSSTH),Bhavnagar | |||
|---|---|---|---|
| Document No.1 | Document Name: Total Cholesterol Examination Procedure | Unique ID:LSSTH /BIOCHEM/ SOP-5 | |
| Issue No. : 01 | Issue Date :30/04/2024 | Page No. | |
| Amend No. | Amend Date | Prepared by: Section Incharge | Approved & Issued by: HOD,Biochemistry |