Immunohistochemistry in human placental tissue--pitfalls of antigen detection.
Because incongruous controversial staining results are a common phenomenon in the placenta, methodical investigations are important to prevent researchers from obtaining misleading results. While investigating dendritic cells (DC) at the human fetomaternal interface, we observed staining of endothelial cells (EC) in chorionic villi for CD83. Given the high specificity of this antigen for DC, this did not seem credible. Previous studies had revealed the same surprising staining pattern with human leukocyte antigen (HLA)-G antibodies. We therefore analyzed human placental EC staining more closely. Both CD83 and HLA-G antibodies were of the same mouse IgG2b isotype. We also observed EC staining with a panel of control antibodies of the IgG2b isotype. This suggests a high affinity of human placental capillaries for mouse IgG2b. Several commonly used techniques for blocking nonspecific binding of antibodies could not prevent this nonspecific EC staining. A new preincubation step with purified human IgG was introduced. This abolished any placental EC staining with CD83, HLA-G, and IgG2b isotype control antibodies, presumably by blocking Fc receptors, whereas specific staining patterns remained unchanged. Mouse antibody of the IgG2b isotype are bound nonspecifically by vascular endothelial cells in human placenta and this can be overcome by blocking with purified human IgG. This blocking procedure could also be appropriate for frozen tissues other than placenta in which Fc receptors are expressed.
Honig
Arnd
A
Department of Obstetrics and Gynecology, Josef-Schneider-Strasse 4, D-97080 Würzburg, Germany.
Rieger
Lorenz
L
Kapp
Michaela
M
Dietl
Johannes
J
Kämmerer
Ulrike
U
D000818:Animals; D000918:Antibody Specificity; D000941:Antigens / Q000032:analysis* / Q000276:immunology; D015703:Antigens, CD / Q000032:analysis / Q000276:immunology; D002824:Chorionic Villi / Q000378:metabolism; D042783:Endothelial Cells / Q000378:metabolism; D005260:Female; D006680:HLA Antigens / Q000032:analysis / Q000276:immunology; D059951:HLA-G Antigens; D015395:Histocompatibility Antigens Class I / Q000032:analysis / Q000276:immunology; D006801:Humans; D007074:Immunoglobulin G; D007136:Immunoglobulins / Q000032:analysis / Q000276:immunology; D007150:Immunohistochemistry / Q000379:methods; D008562:Membrane Glycoproteins / Q000032:analysis / Q000276:immunology; D051379:Mice; D010920:Placenta / Q000098:blood supply / Q000378:metabolism* / Q000648:ultrastructure; D011247:Pregnancy; D011261:Pregnancy Trimester, First; D011961:Receptors, Fc / Q000032:analysis; D000097802:CD83 Antigen
D000941:Antigens; D015703:Antigens, CD; D006680:HLA Antigens; D059951:HLA-G Antigens; D015395:Histocompatibility Antigens Class I; D007074:Immunoglobulin G; D007136:Immunoglobulins; D008562:Membrane Glycoproteins; D011961:Receptors, Fc; D000097802:CD83 Antigen