Production and characterization of monoclonal antibodies directed against the laminin receptor precursor.
The 67-kDa laminin receptor (67LR) is an important tumor marker whose molecular structure has not yet been fully elucidated. To shed new light on this molecule, we raised a series of eight new monoclonal antibodies, designated MPLR1 to 8, directed against the 37-kDa recombinant laminin receptor precursor (37LRP). Cross-competition experiments demonstrated that the epitopes recognized by MPLR2, 4 and 5 partially overlap, since MPLR4 and 5 compete with labelled MPLR2 for the binding to recombinant 37LRP. These three antibodies belong to the IgG1 class, whereas the other ones are all IgM. Presumably due to the fact that they are directed against partially unfolded antigenic determinants expressed on the recombinant protein, MPLRs did not recognize the native protein. Indeed, they showed no reactivity at the membrane level in cytofluorimetric analysis and they did not work in immunoprecipitation experiments. In contrast, these reagents are valuable tools in immunoblotting, since they clearly identify a 67-kDa protein (the mature laminin receptor) in addition to the 37-kDa precursor form. MPLRs are thus a new powerful tool which could help in the characterization of the still enigmatic 67LR molecule.
Butò
S
S
Division of Experimental Oncology E, Istituto Nazionale Tumori, Milano, Italy.
Ghirelli
C
C
Aiello
P
P
Tagliabue
E
E
Ardini
E
E
Magnifico
A
A
Montuori
N
N
Sobel
M E
ME
Colnaghi
M I
MI
Ménard
S
S
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000276:immunology*; D001667:Binding, Competitive; D014408:Biomarkers, Tumor / Q000276:immunology*; D015153:Blotting, Western; D000939:Epitopes / Q000276:immunology; D005434:Flow Cytometry; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D006825:Hybridomas / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D011498:Protein Precursors / Q000276:immunology*; D017474:Receptors, Laminin; D011994:Recombinant Proteins / Q000276:immunology; D012269:Ribosomal Proteins / Q000276:immunology*; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D014408:Biomarkers, Tumor; D000939:Epitopes; D011498:Protein Precursors; D017474:Receptors, Laminin; D011994:Recombinant Proteins; D012269:Ribosomal Proteins