Birkenhäger-Frenkel D H, Birkenhäger J C
Bone. 1987;8(1):7-12. doi: 10.1016/8756-3282(87)90125-6.
On theoretical grounds, it seemed plausible that the apparent distance between two fluorescent labels can surpass the actual distance between them by a percentage that is related to the true distance: section thickness ratio. For similar reasons, the percentage of double labels discernible as such would be influenced by the same ratio. These assumptions were tested in a series of doubly labeled biopsies in which a large variation of appositional rates had been found. The apparent mean distance between labels and, hence, the appositional rate as measured in 20 microns sections was significantly lower than in the 5 microns sections from the same biopsies. Yet, 20 microns sections cannot be used to counteract distortion by oblique sectioning because of the bias obscuring the lower range of true interlabel distances. The percentage of doubly labeled surfaces was from 13 to over 50% lower in the 20 microns sections. Thus, overlap of double labels by geometrical projection in 20 microns sections causes a considerable reduction of formation rates derived from the parameters investigated. The problem is resolved, to a large extent, by using sections as thin as possible.
从理论上讲，两个荧光标记之间的表观距离超过它们实际距离的百分比似乎与真实距离：切片厚度比有关，这似乎是合理的。出于类似的原因，可识别的双重标记的百分比也会受到相同比例的影响。在一系列双重标记的活检组织中对这些假设进行了检验，在这些活检组织中发现了很大范围的贴壁率变化。在相同活检组织的20微米切片中测量的标记之间的表观平均距离以及由此得出的贴壁率明显低于5微米切片中的。然而，由于偏差会掩盖较低范围的真实标记间距离，20微米切片不能用于抵消斜切造成的失真。在20微米切片中，双重标记表面的百分比要低13%到50%以上。因此，20微米切片中通过几何投影造成的双重标记重叠会导致从所研究参数得出的形成率大幅降低。通过使用尽可能薄的切片，在很大程度上解决了这个问题。
