利用三維折射率顯微術應用於定量式分析海洋性貧血

Abstract

生活中貧血是常見的人體疾病，在輕度時病患並不會有感覺到異常，有時也會伴隨其他疾病而發生，貧血是由於血液的攜氧效率不足所致，而血液的攜氧效率與紅血球的形態有關，包含紅血球體積、形狀等，臨床對於懷疑有貧血的病患第一步就是抽血檢驗，並利用全自動分析儀計算出紅血球的相關數值，供醫師做診斷。 貧血有許多種類別，例如：缺鐵性貧血、海洋性貧血、再生不良性貧血、骨髓化生不良症等等，不同的類型有不同的診斷依據，在本篇研究當中，著重於海洋性貧血，海洋性貧血是由於基因變異導致紅血球體積小及血紅素含量低落，其所影響的因素較為單一，與血小板、白血球的多寡影響無關，故選用此疾病作為研究主題。 臨床所使用的儀器是採用阻抗計數及流式細胞儀分析，此儀器的優勢是可以提供大量且快速的檢驗結果，信息量大，但缺點是無法比較單一血球間的差異，因其所得資訊是利用電學所換算的資訊無法觀察到血球型態，且儀器價格及維護費昂貴，故全自動分析儀可以作為臨床初步的診斷，若要更進一步做分析，仍需人工處理染色分析。基於本實驗室所開發的三維折射率顯微術的應用，藉由干涉影像量測樣本完整資訊，再透過光學繞射斷層演算法，類似於電腦斷層的掃描概念重建出三維影像，並對單一血球做幾何量化分析，最後再比較不同顆之間的數值差異，包含總相位、體積、表面積、球型指數、血紅素含量、血球內血紅素濃度、折射率、折射率增益值，其中折射率增益值會透過不同濃度之人體萃取血紅素溶液後，打入微流道量測而得，結果顯示海洋性貧血之折射率增益值標準差較正常人大，其他幾何參數及量化參數則皆有統計上顯著性差異。

Hematological properties of erythrocytes are associated with various blood-related diseases. Clinically, physicians make a diagnosis mainly based on blood reports obtained using Complete Blood Count (CBC), which is a common tool to measure average values of fundamental indicators such as the volume, hemoglobin content and hemoglobin concentration of all erythrocytes analyzed. However, the information about individual erythrocytes, as well as morphological information, is unavailable for physicians. Common-path tomographic diffractive microscope (cTDM) is a novel three-dimensional (3-D) quantitative phase imaging technique that can acquire 3-D refractive index (RI) images of living cells. We employed cTDM to acquire 3-D RI images of erythrocytes obtained from normal volunteers and patients with mild thalassemia. Only about 0.5 μL of blood is sufficient for imaging with cTDM while CBC needs at least 1 mL. Additionally, we calculated the dry mass of each erythrocyte from phase images after experimentally estimating the specific refractive increment of hemoglobin using custom-made microfluidic channels. The diagnostic accuracy of various indices demonstrates that the total phase and the ratio of volume to surface area are the best parameters to distinguish between the normal and thalassemia erythrocytes.