利用三倍頻顯微術量化活體內黑色素質量密度

In Vivo Quantification of Melanin Mass Density by Using Third Harmonic Generation Microscopy

劉威民(Wei-Min Liu)

指導教授 ： 孫啟光

國立臺灣大學/電機資訊學院/光電工程學研究所/碩士(2013年)

https://doi.org/10.6342/NTU.2013.02695

摘要

我們以鉻貴橄欖石鎖模鐳射（中心波長一千兩百三十奈米坐落於皮膚內的吸收和散射較少的穿透窗口）實現活體內虛擬切片。於先前的研究成果中發現，黑色素提供了強烈的三倍頻對比，並隨著膚色越深而更增強。在此研究中，更加深入探討此增益效應。運用老鼠黑色素瘤細胞作為模型，並以二光子螢光訊號作為黑色素濃度的依據，同時取得二光子螢光和三倍頻訊號，進以分析其關係。我們根據二光子螢光頻譜，選用適當的黑色素及溶劑做密度校正。結果顯示，黑色素質量密度和二光子螢光呈線性關係，進而聯結了三倍頻訊號增益與黑色素質量密度。藉由白斑病病人的影像作校正後，找到了適當的比較基準。在集合相關文獻比對之後，也確認了此技術的預測範圍坐落在合理的區塊。最終實現了活體內定量黑色素質量密度，而這也是現階段世界上唯一同時擁有次微米解析度和絕對黑色素質量密度定量的活體內觀測技術。

關鍵字

三倍頻顯微術 ； 活體內 ； 黑色素 ； 質量密度 ； 定量

並列摘要

In previous research, one realized in vivo virtual biopsy using higher harmonic generation microscopy based on a Cr:forsterite laser. Melanin is proved to provide strong contrasts, and the strength of these contrasts enhanced with skin type. In this study, I investigate this enhancement phenomenon. I utilized B16-F10 melanoma cell as the biological model to study in vivo melanin induced third-harmonic generation (THG) enhancement. I used the endogenous two-photon-excited fluorescence (TPEF) as the concentration indicator, and a further calibration between fluorescence and melanin mass density (MMD) had been conducted. I found a good linear relation between TPEF and MMD with R2 equal to 0.98. I took simultaneous THG and TPEF images, and the THG intensity was transformed into THG enhanced ratio by carefully selection of background THG of cytoplasm and TPEF could be transformed into MMD. The relation between the THG enhanced ratio and MMD was therefore constructed. For in vivo HG images, I found that the THG strength of basal cytoplasm without melanin is very similar to that of papillary dermis (PD). For this reason, I could use the THG of PD as the comparison standard to calculate the THG enhanced ratio. By comparing our estimation result with that derived from literature, I can verify that our result lies in a reasonable range. Finally, the first in vivo melanin mass density quantification with a submicron resolution and absolute values of melanin quantity had been realized.

並列關鍵字

Third harmonic generation microscopy ； In vivo ； Melanin ； Mass density ； Quantification

參考文獻

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