Horseradish peroxidase (HRP) conjugates of cholera toxin and lectins are more sensitive retrogradely transported markers than free HRP.

Horseradish peroxidase (HRP) conjugates of 8 different lectins (wheat germ agglutinin, ricinus communis I and II, peanut agglutinin, lens culinaris, soybean agglutinin, limulus polyhemus, ulex europaeus I) and cholera toxin (CT) or free HRP (FHRP) were individually injected into the submandibular gland (SMG) or anterior chamber (AC) of the eye and the retrogradely labeled neurons in the superior cervical ganglion (SCG) were quantitated. The effect of using 3 different cross-linking reagents (glutaraldehyde, p-benzoquinone and periodic acid) on the results obtained with HRP conjugates of wheat germ agglutinin (WG) was also examined. The results in 100 rats demonstrated the superior sensitivity of ligand-HRP conjugates over FHRP as retrogradely transported markers. After SMG injections, HRP conjugates of CT, WG and soybean agglutinin were 20-50 times more sensitive than FHRP. After AC injections, HRP conjugates of CT and WG consistently yielded labeled SCG neurons while FHRP failed to do so even when the amount of FHRP injected was increased 10-fold. The sensitivity of HRP conjugates of WG was similar after SMG injections using each of the 3 cross-linking reagents, but AC injections of conjugates produced with p-benzoquinone yielded twice as many labeled SCG neurons as the other WG conjugates. Mixtures of conjugates with and without FHRP were no more sensitive than the most sensitive individual ligand-HRP conjugates, except after SMG injections of a conjugate mixture with FHRP. Additional experiments demonstrated the specificity of the ligand-"receptor" interaction of WG and CT and that the superior sensitivity of these ligand-HRP conjugates does not depend on the tissue destruction produced by the injection procedure.