Nuclear localization of antigens detected by a monoclonal antibody against a synthetic peptide of rfp finger protein.
A monoclonal antibody (MAb) was generated against a synthetic peptide corresponding to amino acids 148 to 163 of the rfp protein with zinc finger domains. The MAb, designated RFP-1 (IgM), which was positive with the immunizing peptide in enzyme-linked immunosorbent assay, was reactive in immunoblotting with an in vitro translated rfp product as well as with native proteins in cell extracts made from mouse testis and HL-60 human leukemia cell line, both of which were previously shown to express high levels of rfp mRNA. When HL-60 cells were fractionated into nuclear and cytoplasmic components, the protein reactive with RFP-1 MAb was detectable only in the nuclear fraction. By the avidin-biotin complex immunoperoxidase method, this MAb strongly stained over 90% of the nuclei of human and mouse spermatogenic cells, except mature spermatozoon, and of human testicular tumor cells. In other human adult tissues, up to 60% of positive cells were observed. These antibody activities were clearly absorbed by pre-incubation of RFP-1 MAb with the immunizing peptide. These results thus indicated that RFP-1 MAb recognizes a nuclear protein which is expressed at high levels in male germ cells.
Kikuchi
T
T
Department of Immunology, Nagoya University School of Medicine, Japan.
Takahashi
M
M
Ueda
R
R
Ohba
M
M
Seito
T
T
Hiai
H
H
Nakashima
I
I
Takahashi
T
T
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D000918:Antibody Specificity; D015153:Blotting, Western; D002467:Cell Nucleus / Q000032:analysis*; D004268:DNA-Binding Proteins / Q000032:analysis*; D006801:Humans; D007150:Immunohistochemistry; D008667:Metalloproteins / Q000032:analysis*; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000032:analysis*; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000276:immunology; D011994:Recombinant Proteins / Q000032:analysis; D044767:Ubiquitin-Protein Ligases; D000070584:Tripartite Motif Proteins
D000911:Antibodies, Monoclonal; D004268:DNA-Binding Proteins; D008667:Metalloproteins; D009687:Nuclear Proteins; D010446:Peptide Fragments; D011994:Recombinant Proteins; D044767:Ubiquitin-Protein Ligases; C064419:TRIM27 protein, human; D000070584:Tripartite Motif Proteins; C489296:Trim27 protein, mouse