Hybridomas producing monoclonal anti-idiotypic antibodies (anti-id MAbs) to N-acetylglucosaminyl-B1-4-N_acetylmur~yl-~~yl-~-isoglutamine (GMDP) were developed. Three clones of hybridomas demonstrated the properties characteristic for the Ab2# type of anti-id antibody: they bound to Fab-fragments of high-affinity MAb to GMDP, dosedependent inhibition of this binding by GMDP was observed; immunization of mice with these MAbs resulted in production of GMDP-specific antibodies. When these antibodies were used to stain blots from SDS-PAGE of macrophage lysate, the same receptor proteins were specifically stained as upon staining with 'zsI-labelled GMDP derivative. Muramyldipeptide(MDP) is the minimal structure of the bacterial cell wall capable of replacing mycobacteria in Freunds' complete adjuvant [l]. MDP and its analogues, containing the N-acetylglucosamine residue, demonstrate a variety of biological activities, in particular, adjuvant effect [2,3], induction of non-specific resistance to infections [4,5], and som-.nogenic activity [6]. The molecular basis of biological activity of mummy1 peptides (MPs) is poorly understood, but the majority of data favors a receptor-mediated mechanism [7-lo].Macrophages are shown to be the main target of MPs [4]. The number of cell surface receptors for MPs is low, making their isolation and structural analysis difficult [7,8]. Like in studies of receptors of other biologically active ligands, monoclonal antibodies to GMDP-receptors could be of great aid, but their production by traditional procedures is hardly achieved.Not long ago an alternative approach was developed based on the production of anti-idiotypic antibodies (anti-id) to corresponding ligands, capable of binding to ligand receptors. This approach is based on the Jeme theory of idiotypic network, claiming that the immune system consists of a set of interacting idiotypes and anti-idiotypes [ 111. According to this theory for each exogeneous epitope an anti-id antibody exists mimicking the so-called internal image of this epitope by its hypervariable region. This antibody is designated as Ab2/3 [12].Anti-id antibodies can be induced by immunization with anti-ligand antibodies, imitating the binding site of the receptor. Using this approach monoclonal and polyclonal antibodies to a number of receptors were produced [13-161. The crucial step in obtaining anti-id MAbs capable of binding to the ligand *Corresponding author. Fax: (7) (095) 310 7007.Abbreviations: MP, mummy1 peptide; MDP, MurNAc-Ala-n-iGln; GMDP, GlcNAc-/314MurNAc-Ala-D-iGln; L-GMDP, GlcNAc#?1-4-MurNAc-Ala-iGin; GMDP-Lys, GlcNAc-/714-MurNAoAla-n-iGlnLys; PBS, 0.01 M sodium phosphate buffer, containing 0.15 M NaCl; PBST, PBS containing 0.1% Tween 20; BSA, bovine serum albumin; GMDP-OVA, GMDP conjugated to ovalbumin; GMDP-AL, GMDP conjugated to poly-Lys-poly-Ala polymer; MeBSA, methylated bovine serum albumin.receptor is identifying among hybridoma those clones producing antibodies of the Ab2# type to paratope-associated epitopes. In this study we ...