Diagnosis of Crystalopathies via Raman Spectroscopy

描述（由申请人提供）：关节间隙中晶体种类的存在和识别的误诊通常会因关节病变条件的错误治疗而受到惩罚。现有的偏振成像诊断标准缺乏特异性和敏感性，主要局限于显微镜下可见的双折射晶体。在目前的研究中，我们提出拉曼光谱作为一种替代的诊断技术，以改进关节空间中晶体种类的识别。在拉曼光谱学中，分子振动是通过激光激发触发的，反射光保存着观察区域中物种的身份和数量的信息。该技术目前用于其他疾病的关节镜体内诊断；因此，基于这种临床潜力，探索其在关节病变条件下晶体种类评估的分析和临床可行性是值得的。提出的探索性研究的第一阶段将揭示各种晶体的识别阈值，可以通过拉曼光谱检测。将合成的尿酸钠、二水合焦磷酸钙、羟基磷灰石、磷酸八钙、磷酸三钙、草酸钙、胆固醇、胱氨酸和黄嘌呤等类似物与不同浓度的健康滑液样本混合，并在宏观水平上分析这些混合物，以检测晶体发出的累积信号。我们将尝试通过采用表面增强拉曼光谱和光谱数据的主成分分析来降低识别阈值。在第二阶段，将从患者身上采集滑液样本，并通过补偿偏振显微镜、x射线衍射和茜素染色来确认晶体的存在或不存在。这些样品的等分将进行拉曼分析，以确定拉曼分析是否可以检测到临床样品中晶体的存在和身份。如果这项探索性研究表明拉曼分析的分析极限足以覆盖晶体种类的临床浓度，那么可以进行全面的临床研究来评估该技术的特异性和敏感性，最终导致关节镜拉曼装置的发展。

DESCRIPTION (provided by applicant): Misdiagnosis of the presence and the identity of crystal species in joint space are often penalized with the incorrect treatment of the arthropathic condition. The existing diagnostic standard of polarized imaging lacks specificity and sensitivity and it is largely limited to microscopically visible birefringent crystals. In the current study we propose Raman spectroscopy as an alternative for diagnostic technology for improved identification of crystal species in joint spaces. In Raman spectroscopy molecular vibrations are triggered through laser excitation and the reflected light holds information on the identity as well as on the amount of species in the observed region. The technique is currently utilized for arthroscopic in vivo diagnosis of other diseases; thus, based on this clinical potential it would be worthwhile to explore its analytical and clinical feasibility towards the assessment of crystal species involved in arthropathic conditions. The first phase of the proposed exploratory study will reveal the identification thresholds of various crystals that can be detected by Raman spectroscopy. Synthetic analogs of monosodium urate, calcium pyrophosphate dihydrate, hydroxyapatite, octacalcium phosphate, tricalcium phosphate, calcium oxalate, cholesterol, cystine and xanthine will be mixed with healthy synovial fluid samples at various concentrations and these mixtures will be analyzed at the macro level to detect the cumulative signal emanating from crystals. We will attempt to lower the identification thresholds by employing surf ace-enhanced Raman spectroscopy and by principal component analysis of the spectral data. During the second phase synovial fluid samples will be taken from patients and the presence or absence of crystals will be confirmed via compensated polarized microscopy, X-ray diffraction and alizarin staining. Aliquots of these samples will be subjected to Raman analysis to determine whether Raman analysis can detect the presence and identity of crystals in clinical samples. If this exploratory study demonstrates that the analytical limits of Raman analysis suffice to cover clinical concentrations of crystal species than comprehensive clinical studies can be conducted to assess the specificity and the sensitivity of the technique finally leading to the development of an arthroscopic Raman device.