Lj. Freedman et Mt. Maddox, A comparison of anti-biotin and biotinylated anti-avidin double-bridge andbiotinylated tyramide immunohistochemical amplification, J NEUROSC M, 112(1), 2001, pp. 43-49
Often it is difficult to detect very small amounts of antigen with conventi
onal immunohistochemical techniques. We evaluate three amplification techni
ques involving anti-biotin or anti-avidin double-bridges or biotinylated ty
ramide amplification to enhance the sensitivity of serotonin transporter im
munohistochemistry. For the anti-biotin double-bridge, after the secondary
antibody, the sections were incubated in anti-biotin antibody followed by a
second incubation in the secondary antibody and then avidin-biotin-peroxid
ase complex (ABC). For the biotinylated anti-avidin technique, after the AB
C, sections were incubated in biotinylated anti-avidin, followed by another
incubation in ABC. For the biotinylated tyramide technique, after the ABC
step, sections were incubated in biotinylated tyramide and hydrogen peroxid
e, followed by another incubation in ABC. The anti-biotin double-bridge als
o resulted in a large increase in the number of stained fibers and the inte
nsity of labeling with no increase in background. A biotinylated anti-avidi
n double-bridge also produced significant signal amplification but signific
ant background. The biotinylated tyramide technique resulted in an even lar
ger increase in the number of labeled fibers and an intensity of their stai
ning with a moderate amount of background staining. However, this advantage
was not present at high dilutions of primary antibody. Thus, the anti-biot
in double-bridge is likely to be useful in immunohistochemistry and immunof
luorescence as well as other situations where increased sensitivity and low
background from biotin markers is needed. The biotinylated tyramide techni
que may also be useful where some degree of background labeling is acceptab
le. (C) 2001 Elsevier Science B.V. All rights reserved.