Miksche J P, Dhillon S S, Berlyn G P, Landauer K J
J Histochem Cytochem. 1979 Oct;27(10):1377-9. doi: 10.1177/27.10.92496.
Nonspecific light loss by the cell-wall-plus-cytoplasm (CWC) can cause a 50% increase in Feulgen absorption units in peanut root-tip nuclei as determined by scanning at 450 nm, whereas this phenomenon is not evident with chicken erythrocytes. A two wavelength scanning method of subtracting nonspecific 450 nm absorption from 550 nm Feulgen absorption values eliminated the nonspecific light loss in CWC, However, the two wavelength scanning method is time consuming and somewhat impractical with a regular scanning microdensitometer such as Vickers M85. Elimination of the problem of nonspecific light loss is suggested by careful determination of background setting with the spot position close to the nucleus in CWC. The accuracy of the CWC background setting method was further tested by comparison with subtraction method. The use of plant nucleis as an internal standard in plant DNA measurements was also evaluated. Significant variation among the replicate slides due to the variation in pine nuclear DNA amounts was observed and plant nuclei generally are not reliable internal standards. Mature chicken erythrocytes are recommended as an internal standard because the cell type and metabolic state is known.
细胞壁加细胞质（CWC）造成的非特异性光损失可使花生根尖细胞核中的福尔根吸收单位增加50%，这是通过在450nm处扫描测定的，而鸡红细胞则未出现这种现象。一种从550nm福尔根吸收值中减去450nm非特异性吸收的双波长扫描方法消除了CWC中的非特异性光损失。然而，双波长扫描方法耗时，对于像维氏M85这样的常规扫描显微密度计来说有些不切实际。通过仔细测定CWC中靠近细胞核的光斑位置的背景设置，建议消除非特异性光损失问题。通过与减法方法比较，进一步测试了CWC背景设置方法的准确性。还评估了在植物DNA测量中使用植物细胞核作为内标。观察到由于松树核DNA量的变化，重复玻片之间存在显著差异，并且植物细胞核通常不是可靠的内标。建议使用成熟鸡红细胞作为内标，因为细胞类型和代谢状态是已知的。
