Murine monoclonal anti-idiotype antibody (alpha) as a probe to detect human monoclonal antibody bound to human tumor tissues.
A new immunohistochemical assay was developed for the detection of human monoclonal antibody (HuMAb) bound to human biopsied tumor tissues. A murine anti-idiotype monoclonal antibody, alpha type, 18C6 (IgGl), was raised against an IgM HuMAb, L612, defining a tumor-associated ganglioside antigen (GM3) and used as a probe in a three step cell-binding assay (HuMAb + anti-id + biotinylated anti-mouse Ig). Anti-id 18C6 has an exclusive binding specificity for HuMAb L612, but does not interfere with the binding of L612 to antigen positive melanoma cell lines or to a purified antigen, GM3. The applicability of 18C6 in the three step cell-binding assay was tested first using a melanoma cell line, UCLASO-M12. L612 bound to M12 cells was specifically detected by 18C6 without any background reactivity in ELISA. When this assay was compared with the standard two-step cell-binding assay (HuMAb + peroxidase-conjugated anti-human IgM) using various cultured tumor cell lines, parallel reactivity was observed. The three-step cell-binding assay was then applied to various fresh-frozen human tumor sections. Positive reactivity was demonstrated on various histologic types of human tumor tissues: primary melanoma (10/10), metastatic melanoma (4/4), nevus (10/10), lung cancer (3/6), breast cancer (2/6), and colon cancer (1/1). Adjacent normal tissues were unstained. Control experiments included the cell-binding assay with L612 alone, 18C6 alone. L612 + unrelated mouse IgG, and unrelated IgM HuMAb (L72) + 18C6; but biotinylated anti-mouse IgG did not react with these control preparations. The results indicate that anti-id 18C6 is a highly specific probe to assess the expression of the ganglioside antigenic epitope recognized by the L612 HuMAb on biopsied human tumor tissues.
Saito
H
H
Division of Surgical Oncology, UCLA School of Medicine 90024.
Wen
D R
DR
Yamamoto
S
S
Yamamoto
T
T
Saxton
R E
RE
Cochran
A J
AJ
Irie
R F
RF
D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000276:immunology; D000911:Antibodies, Monoclonal / Q000032:analysis* / Q000276:immunology; D001706:Biopsy; D004797:Enzyme-Linked Immunosorbent Assay; D005679:G(M3) Ganglioside / Q000276:immunology; D006801:Humans; D007124:Immunoenzyme Techniques; D007150:Immunohistochemistry / Q000379:methods*; D051379:Mice; D008807:Mice, Inbred BALB C; D009369:Neoplasms / Q000276:immunology* / Q000473:pathology; D012680:Sensitivity and Specificity; D014407:Tumor Cells, Cultured / Q000276:immunology
D000888:Antibodies, Anti-Idiotypic; D000911:Antibodies, Monoclonal; D005679:G(M3) Ganglioside