Simple immunohistochemical staining method using large sized gold colloid conjugated secondary antibody.
To test the feasibility of semiquantitative immunohistochemical staining (IHC) with large sized gold conjugated secondary antibody (gold-2 degrees Ab), we used beads covered with a known amount of primary antibody and a secondary antibody conjugated with gold colloid particles (20 and 40 nm in diameter), and we compared the results to those obtained by enzyme IHC. Beads coated with 6 graded amounts of mouse IgG molecules showed 6 levels of color intensity. The graded color intensities could readily be distinguished. The color developed as soon as we added gold-2 degrees Ab, and the intensities were stable for 1 wk. Enzyme IHC using identical beads showed dregs of pigment after incubation in DAB for 5 min. The large sized gold-2 degrees Ab showed strong signals on cell surfaces; application of the large sized gold-2 degrees Ab to paraffin-embedded tissue sections was also feasible. The color was bright red and was easier to differentiate from hemosiderin pigment than the color developed by enzyme IHC. In conclusion, gold IHC with large sized gold-2 degrees Ab is superior to enzyme IHC for quantification of antigens via IHC. Gold IHC is especially recommended for tissues with many macrophages, such as bone marrow and spleen.
Lee
Eun Jung
EJ
Department of Laboratory Medicine, Hospital Pathology, Catholic University Medical College, Seoul, Korea.
Kim
Yonggoo
Y
Lim
Jihyang
J
Kim
Myungshin
M
Kang
Chang Suk
CS
Lee
Jae Hoon
JH
Mok
Rak Sun
RS
Park
Young Nam
YN
Choi
Ha-Young
HY
Han
Kyungja
K
D000818:Animals; D018956:CD5 Antigens / Q000032:analysis; D005240:Feasibility Studies; D019084:Fluorescent Antibody Technique, Indirect / Q000379:methods*; D017979:Gold Colloid / Q000737:chemistry*; D006801:Humans; D018796:Immunoconjugates; D007124:Immunoenzyme Techniques; D007963:Leukocytes, Mononuclear / Q000737:chemistry; D051379:Mice; D010316:Particle Size; D051381:Rats; D013194:Staining and Labeling
D018956:CD5 Antigens; D017979:Gold Colloid; D018796:Immunoconjugates