CD95 ligand (CD95L) immunohistochemistry: a critical study on 12 antibodies.
In recent years, some studies on the expression of CD95(Fas/APO-1) ligand (CD95L) in tissues or cells raised concerns about the specificity of the antibodies used. We therefore tested 12 CD95L antibodies for their reliability in immunocyto/histochemistry by (i) staining CD95L-transfected and control CV-1/EBNA cells and (ii) comparing staining patterns in immunohistochemically labeled tissue sections with the localization of CD95L+ cells in in situ hybridization. While G247-4, NOK-1, NOK-2, 4H9, and MIKE-1 stained CD95L-transfected cells and did not significantly bind to controls, G247-4 was the only antibody giving satisfying signals in tissue sections perfectly matching the distribution of CD95L+ cells by in situ hybridization. MAb 33, C-20, and N-20 comparably stained both transfected and control cells and showed considerable background or falsely positive staining in sections. MIKE-2, 8B8, A11, and 4A5 did not or only very faintly bind to either cells and, thus, were not tested on sections. We conclude that G247-4 is the only tested antibody that is recommendable for immunohistochemistry.
Sträter
J
J
Institute of Pathology, University of Ulm, Albert-Einstein-Allee 11, D-89081 Ulm, Germany. joern.straeter@medizin.uni-ulm.de
Walczak
H
H
Hasel
C
C
Melzner
I
I
Leithäuser
F
F
Möller
P
P
D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D051017:Apoptosis Regulatory Proteins / Q000096:biosynthesis; D053222:Fas Ligand Protein; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D017403:In Situ Hybridization; D008562:Membrane Glycoproteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D014066:Palatine Tonsil / Q000737:chemistry / Q000276:immunology; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction; D053221:TNF-Related Apoptosis-Inducing Ligand; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis; D048069:Tumor Necrosis Factors / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*
D000911:Antibodies, Monoclonal; D051017:Apoptosis Regulatory Proteins; C505661:FASLG protein, human; D053222:Fas Ligand Protein; D008562:Membrane Glycoproteins; D012333:RNA, Messenger; D053221:TNF-Related Apoptosis-Inducing Ligand; C505653:TNFSF10 protein, human; D014409:Tumor Necrosis Factor-alpha; D048069:Tumor Necrosis Factors