A10%solutionofFluoro-Rubyismadebydissolving10mgofFluoro-Rubydrypowderin100µLofdistilledwater.Itisthendeliveredviaintracranialstereotaxicinjectionusinga1uLHamiltonmicrosyringetopressureinjectthetracer.Injectionvolumestypicallyrangefrom0.02-0.1µLandaregraduallyinjectedovera10-15minuteinterval.Theanimalisallowedtorecoverandissacrificedtypically4-14dayslater.

Theanimalisthenperfusedwithneutralbufferedformaldehyde(10%formalinor4%paraformaldehydein0.1Mneutralphosphatebuffer.)Thebrainisremovedandpostfixedforatleastovernightinthesamefixativesolutionplus20%sucroseforcryo-protection.Thebrainisthencutonafreezingslidingmicrotomeorcryostatintosectionsusuallybetween20and40micronsinthickness.

Thetissuesectionsaremountedongelatincoatedslidesandareairdriedonaslidewarmerat50-60degreesCentigradeforatleastonehalfhour.ThedryslidesarethentransferredtoxyleneclearingsolutionforatleastoneminuteandthencoverslippedwithDPXmountingmedia.TheslidescanthenbeexaminedunderanepifluorescentmicroscopeusingafiltersuitableforvisualizingTRITC(greenlightexcitation).

DAPI染色的纵截面显微照片，通过小鼠右颈脊髓，显示了荧光金标记的运动神经元的典型列。该图最初发表于Tosolini AP，Mohan R，Morris R（2013）。靶向小鼠前肢中运动终板区域的全长可增加氟金对相应脊髓运动神经元的吸收。面前。神经元。4：58、1-10、4

大鼠前肢强荧光荧光金标记的运动神经元。该图最初发表于Tosolini AP，Mohan R（2012）。供应大鼠前肢的运动神经元柱的空间表征。神经科学200：19-30，22。

成年小鼠腰脊髓的显微照片，显示了用DAPI复染色的典型的荧光金标记柱。该图最初发表于Mohan R，Tosolini AP，Morris，R.（2014）。神经科学274：318-330，323