Veh R W
Abteilung für Neuroanatomie, Ruhr-Universität Bochum, F.R.G.
Neurosci Lett. 1991 Jan 2;121(1-2):169-72. doi: 10.1016/0304-3940(91)90677-l.
For double tracing experiments, wheat germ agglutinin (WGA) molecules labeled with two different haptens are desirable. In the present report the suitability of digoxigenylated WGA (DIG-WGA) for retrograde tracing was investigated. For this purpose the new tracer was pressure injected into rat brains and the transported DIG-WGA visualized via its digoxigenyl group with an alkaline phosphatase linked anti DIG antibody in permanently stained sections of high quality. With fixatives containing 2.5% glutaraldehyde only few positive cells were found. However, at milder fixation conditions (4% paraformaldehyde, 0.05% glutaraldehyde 0.2% picric acid, 30 min) retrogradely labeled cells were detected with a sensitivity comparable to tetramethylbenzidine protocols for conventional WGA-HRP (horseradish peroxidase) tracing. Preliminary experiments suggest excellent suitability for double labeling.
对于双重示踪实验，需要用两种不同半抗原标记的麦胚凝集素（WGA）分子。在本报告中，研究了地高辛标记的WGA（DIG-WGA）用于逆行示踪的适用性。为此，将这种新型示踪剂压力注射到大鼠脑中，并在高质量的永久染色切片中，通过其地高辛基团，用碱性磷酸酶连接的抗地高辛抗体使转运的DIG-WGA可视化。仅使用含2.5%戊二醛的固定剂时，发现的阳性细胞很少。然而，在较温和的固定条件下（4%多聚甲醛、0.05%戊二醛、0.2%苦味酸，30分钟），检测到的逆行标记细胞的灵敏度与用于传统WGA-HRP（辣根过氧化物酶）示踪的四甲基联苯胺方法相当。初步实验表明其非常适合双重标记。
