YIELD SINGLE MOLECULE IN FCS COLLECTION VOLUME BY NUMERICAL SIMULATION

我们正在研究多光子自发荧光的用途 组织成像和光谱学，用于基础生物学研究和 作为病理学家潜在的诊断工具。内在组织 荧光可以潜在地提供关于 组织的结构和化学成分。多光子激发 具有更高的激发内在荧光团的潜力 由于排放分离可能非常有效(激励距离很远 从发射区移除)，并且该技术具有 本征光学切片在散射组织中很重要。什么时候 在690-750 nm激发下进行多光子显微镜 区域，组织自发荧光在样品倍数下变得普遍 10-15 mW(约为2 PM功率的3倍) 常规荧光团)。这种方法的明显优势是 组织可能没有任何固定或染色 手工艺品。染料对厚样品的渗透性往往很差。 而固定和脱水技术与约80%的 收缩，一种结构关系没有 必须保存下来。利用自体荧光的缺点 天然的荧光团显然没有针对它们的 荧光特性。因此，仪器配置的优化 对于获取良好的自发荧光图像来说是必不可少的。

We are investigating the uses of multiphoton autofluorescence imaging and spectroscopy in tissue for basic biological research and as a potential diagnostic tool for pathologists. Intrinsic tissue fluorescence can potentially provide significant information about the structural and chemical composition of tissue. Multiphoton excitation has an increased potential for excitation of intrinsic fluorophores since emission separation can be very efficient (excitation is far removed from the emission region) and the technique possesses an intrinsic optical sectioning important in scattering tissues. When performing multiphoton microscopy with excitation in the 690-750nm region, tissue autofluorescence becomes prevalent at specimen powers of 10-15mW (~3-fold higher powers than those used for 2PM of conventional fluorophores). The clear advantage of this method is that the tissue is potentially free from any fixation or staining artifacts. Dye penetration through a thick sample is often poor and fixation and dehydration techniques are associated with an ~80% shrinkage, a collapse in which structural relationships are not necessarily preserved. The disadvantage of utilizing autofluorescence is that natural fluorophores are clearly not optimized for their fluorescent properties. Optimization of instrumentation is thus essential for the acquisition of good autofluorescence images.