Localization of [¹⁴C] Labeled Anti-Streptococcal Cell Membrane Monoclonal Antibodies (anti-SCM mAb) in Mice

Abstract

Six different hybridomas secreting anti-SCM mAb and one control mAb were placed into adult mice along with [¹⁴C] amino acids for biosynthetically labeling. After sacrifice, the ¹⁴C mAb ascites along with serum, heart, kidney, lung and skeletal muscle were recovered. Tissue associated specific radio-activity (SpAc) and microscopic structural analyses were performed. Confirmation of mAb specificity was by both immunodot blots as well as Western blot analysis. Peritoneal injection of measured doses of anti-SCM mAb yielded tissue SpAc confirming their in vitro specificities¹⁰. Two of the mAb showed strong reactivity to both renal (GBM) and lung basement membrane (LBM), two were mainly GBM reactive and two showed polyreactivity with a marked reactivity to a Z-line antigen. Autoradiographic light microscopy confirmed that the anti-SCM mAbs bound to both GBM and LBM and to Z-line antigen. Titrated doses of the mAb yielded autoradiographic confirmation in which the grain number on the GBM and LBM increased with increasing dose of mAb for all mAb except the control. This effect was not seen in the muscle tissues but anatomical localization at the Z-line was consistent. The major significance of these studies is the demonstration that circulating antibodies to SCM can react in vivo with normal mammalian antigens adding confirmation to the in vitro specificity of these cross-reactive anti-SCM mAbs.

Key Words:

• [¹⁴C] mAbs
• Streptococcal membrane
• Cross-reactivity
• Tissue-binding
• Basement membrane
• alpha-actinin
• Z-line