WS/T 617-2018 in English
VALIDReference procedure for the measurement of catalytic activity concentration of aspartate aminotransferase
- Issued on:2018-08-20
- Implemented on:2019-03-01
- File Format:PDF
- Delivery:Via email within 5 business days
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本标准规定了在临床医学应用中，测量天门冬氨酸氨基转移酶催化活性浓度的参考测量程序。
本标准主要适用于参考实验室，作为天门冬氨酸氨基转移酶催化活性浓度测量的溯源，也可作为与酶催化活性浓度检验有关的仪器和试剂生产企业的溯源，可供有关认可监督管理单位及质量管理部门应用。此参考测量程序可用于我国临床常规方法测量结果的溯源。
Introduction
Standard Overview
This standard (WS/T 617-2018) specifies the reference measurement procedure for the catalytic activity concentration of aspartate aminotransferase (AST). It adopts the coupled reaction principle recommended by IFCC and accurately measures the NADH oxidation rate by spectrophotometry at 37°C. The main technical features include:
| Parameters | Indicators |
|---|---|
| Measurement Principle | L-aspartic acid + α-ketoglutarate → oxaloacetic acid + NADH |
| Wavelength | 339±1nm |
| Temperature Control | 37.0±0.1℃ |
| Linear Range | 0.13-4.13 μkat/L |
Key Technical Elements
1. Reagent System
Core reagents include Tris buffer, L-aspartic acid, NADH, LD/MD coupling enzyme, etc. Critical quality control points:
- P-5-P concentration must be accurate to 0.1mmol/L
- NADH purity must be ≥98%
- Enzyme reagents must be stored in glycerol rather than ammonium sulfate
2. Instrument Calibration
| Instrument Type | Calibration Requirements |
|---|---|
| Spectrophotometer | Wavelength accuracy ±1nm, optical path 10.00±0.01mm |
| Point thermometer | Error ≤0.1℃ at 37℃ |
| pH meter | 7.65±0.05(37℃) |
Measurement process optimization
Typical problem solution: When the reagent blank absorbance changes by >0.002/min, check the NADH stability. Actual case studies show that undegassed reaction solutions can increase the blank value by 23%.
- Degassed the reaction solution (stirred in vacuum at 35℃ for 1h)
- Incubated strictly for 300s to fully saturate the P-5-P
- Delayed for 90s to ensure reaction equilibrium
Uncertainty Analysis
Main sources of uncertainty and control measures:
| Component | Contribution | Control method |
|---|---|---|
| Molar absorptivity | 1% | Using IFCC recommended value (ε=630) |
| Temperature fluctuation | 5.3%/℃ | Dual-channel temperature monitoring |
| pH deviation | 18%/0.05pH | Calibrate pH meter daily |
Implementation recommendations
1. Laboratory preparation
- Establish a reagent batch registration system
- Regularly verify the bandwidth of the spectrophotometer (≤2nm)
- Control the ambient temperature at 20-30℃ and the humidity at 30-70%
2. Standardized operation
The following quality control strategy is recommended:
| Control type | Frequency | Acceptance criteria |
|---|---|---|
| Inter-day precision | Daily | CV≤1.5% |
| RELA comparison | Yearly | |Bias|≤2% |
| Linearity verification | When changing batch numbers | r²≥0.998 |
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