We report the development of a National Institute of Metrology (NIM) hemoglobin A1c (HbA1c) certified reference material (CRM). Each CRM unit contains about 10 μL of hemoglobin. Both hemoglobin and glycated hemoglobin were quantitatively determined by high-performance liquid chromatography (HPLC)–isotope dilution mass spectrometry (IDMS) with synthesized VHLTPE and glycated VHLTPE as standards. The mass fraction of synthesized VHLTPE or glycated VHLTPE was also quantitatively determined by HPLC-IDMS with NIM amino acid CRMs as standards. The homogeneity and stability of the CRMs were examined with a commercial HbA1c analyzer based on the HPLC principle. Fifteen units were randomly selected for homogeneity examination, and statistical analysis showed there was no inhomogeneity. Examination of the stability showed that the CRM was stable for at least 6 months at -80 °C. Uncertainty components of the balance, amino acid purity, hydrolysis and proteolysis efficiency, method reproducibility, homogeneity, and stability were taken into consideration for uncertainty evaluation. The certified value of NIM HbA1c CRM was expressed as the ratio of HbA1c to total hemoglobin in moles, and was (9.6 ± 1.9)% . The CRM can be used as a calibration or validation standard for clinical diagnostics. It is expected to improve the comparability for HbA1c measurement in China.

介绍了一种基于吸光度测定的洗板机校准方法,用洗板机对含有考马斯亮蓝-牛血清白蛋白吸光度溶液标准物质的酶标板进行清洗,然后用检定合格的酶标仪分别测定清洗前后酶标板各孔的吸光度,实现对洗涤残留率、交叉污染率、重复性等计量性能的评价.文章同时对洗涤残留率的不确定度评定方法进行了讨论,有助于提高ELISA分析结果的准确度和精密度,并为洗板机校准规范的制定奠定了基础.

本文简单介绍了GUM和MCM不确定度评定的基本原理,以牛血清白蛋白溶液配制为例,分别采用GUM和MCM方法对化学计量中常用的重量—容量法溶液配制结果的不确定度进行了评定.GUM法得到的溶液浓度配制结果为( 16.90 +0.66) g/L,MCM法得到的溶液浓度配制结果为(16.90+0.65)g/L,两者基本一致.其中牛血清白蛋白标准物质引入的不确定度为主要的不确定度分量,对总不确定度的贡献超过了98％,在这种各个不确定度分量大小不相近的情况下,更适合采用MCM法进行不确定度评价.

We present the development process for National Institute of Metrology (NIM) bovine serum albumin (BSA) certified reference material (CRM). Each CRM unit contains about 200 mg of purified BSA. The moisture, ignition residue, molecular weight, and high-performance liquid chromatography (HPLC) purity were analyzed and mass spectrometry based protein identification was carried out to ensure the material was BSA. Both amino acid based isotope dilution mass spectrometry (IDMS) and a purity deduction method were selected for value assignment. The certified value was the average of the IDMS and the purity deduction result. HPLC purity analysis was used to examine the homogeneity and stability of solid BSA CRM. Fifteen units were selected for between-bottle homogeneity examination and seven subsamples from the same bottle were selected for within-bottle homogeneity examination. Statistics showed the CRM passed both the between-bottle and the within-bottle homogeneity examination. The CRM stability under storage conditions (-20 °C) was tested for 18 months and no trend was observed. Uncertainties from the balance, amino acid purity, hydrolysis, method reproducibility, homogeneity, and stability were taken into account in uncertainty evaluation. The final certified value of NIM BSA CRM is (0.963±0.038) g/g.

<正>一、概述牛血清白蛋白第V组分(BSA)是牛血清中的一种球蛋白,包含583个氨基酸残基,分子量约为66430,等电点为4.7。牛血清白蛋白广泛应用于生化实验中,也是最常用的蛋白质定量的标准。为了保证我国蛋白质含量测定结果的准确、可比与可溯源,中国计量科学研究院率先采用同位素稀释质谱法和纯度扣除法研制了牛血清白蛋白固体国家一级标准物质,定值结果为(0.963±0.038)g/g。但

The domestic and international studies on protein measurement and certified reference material (CRM) development within latest 10 years are reviewed, particularly with emphasis attached to isotope dilution mass spectrometry method for protein CRM value assessment and internatioanl comparasion carried out by bioanalysis working group (BAWG). The domestic and international calibration and measurement capability (CMC) claims for protein content measurement as well as development progress of protein content CRMs are also reviewed. Finally, the prospect of protein content CRM research is presented based on the disscussion on latest BAWG and Biological and Environmental Reference Material (BERM) meetings.