FTIR-HATR诊断β-地中海贫血及其机理研究

为建立简单快速的地中海贫血(简称地贫)诊断方法,探讨了傅里叶红外光谱结合水平衰减全反射(FTIR-HATR)技术用于β-地贫患者的检测及其分子机制.检测了68份正常与3Z份β-重型地贫患者样品,并在群体水平上分析了两者光谱差异变化规律.结果显示:(1)由于血红蛋白量的减少,β-地贫组的光谱强度显著低于正常组;(2)在1 750～1 500 cm-1区间,表征蛋白二级结构的1 652,1 638和1 628 cm-1在β-地贫中有明显降低,而在1 682cm-1处略微增强;(3)在1 350 1 500 cm-1区间,表征磷脂CH3/CH2的1 440,1 453,1 479 cm-1及CO-O-C的1 172和1 161cm-1吸收峰在β-地贫中显著减少;而在1 000～1 200 cm-1区间,表征碳水化合物C-O的1 150 cm-1,2,3-二磷酸甘油酸(DPG)或ATP中0-P=0的1 081和1 053cm-1吸收峰显著升高.统计表明DPG/脂类比在β-重型地贫组中显著高于正常组,并可作为诊断指标,将两者完全区分,避免了复杂数据处理过程,为地贫的光谱诊断提供了坚实的实验基础与理论依据.

FTIR-HATR to Identify β-Thalassemia and Its Mechanism Study

Fourier transform infrared spectroscopy (FTIR)associated with horizontal attenuated total reflectance (HATR)was firstly used to diagnose β-thalassemia patients. With excellent linearity (r=0.997) and reproducibility (RSD＜4%), FTIR-HATR shows an order-of-magnitude increase in IR absorption bands over the single-path transmission FTIR. Based on above, spectra from 37 patients’ and 68 health samples indicated several observable differences in IR vibrational spectra of the Hb lysates between the β-thalassemia major patients and control: (1)Because of decreasing hemoglobin, the peak intensities are obviously lower in β-thalassemia group that is consistent with index from routine hemoglobin diagnosis. (2)In 1 750-1 500 cm-1 region, slight decrease at 1 652 cm-1(α-helix), 1 638 and 1 628 cm-1 bands but mild increase at 1 682 cm-1 all demonstrate structure changes by both Fourier self-deconvolution and second derivative spectra. (3)More importantly, difference spectra substantially demonstrate decreased intensities at 1 440, 1 453, 1 479 cm-1 bands arising from CH2/CH3 deformation vibration of phospholipids but increased intensities at 1 150 cm-1 band originating from C—O stretching vibration of carbohydrate and at 1 081 and 1 053 cm-1 bands that are attributed to 2,3-diphosphoglycerate (DPG)in β-thalassemia major group. Statistical analysis demonstrates significant difference of DPG/phospholipids ratio between two groups. All the samples can be 100% correctly classified into groups on the basis of this ratio. These finding could help understand possible mechanism for diagnosing thalassemia. It makes large-scale screening of thalassemia by FTIR a possibility.