Biomedical subjects

Maria Rossi

Publications and source records attributed to Maria Rossi.

3 recordsLinked to original sources

PubMed2002

Affinity purification of polyclonal antibodies using a new all-D synthetic peptide ligand: comparison with protein A and protein G.

This study investigates the applicability of the protein A Mimetic (PAM) affinity ligand, obtained from the screening of a multimeric combinatorial peptide library, in immunoglobulin isolation from serum. To avoid protease degradation, the ligand has been substituted by its inverso form, named D-PAM, synthesized by replacing all amino acids with the corresponding D derivatives. D-PAM affinity columns, prepared by immobilizing the all-D peptide on the commercially available support Emphaze, were able to capture IgG directly from the serum in a single chromatographic step, with a recovery yield ranging from 60% to 90%, a purity degree higher than 90%, and with a full recovery of antibody activity. Column capacity, determined by applying a large excess of purified IgG to 1 ml bed volume column, was close to 52 mg/ml for bovine IgG, 58 mg/ml for goat IgG, 66 mg/ml for horse IgG, 50 mg/ml for human IgG, 52 mg/ml for mouse IgG, 36 mg/ml for rabbit IgG and 48 mg/ml for sheep IgG. D-PAM peptide was found to be very stable to protease activity, and after prolonged incubation with mouse serum. Similarly, the corresponding derivatized matrix tested before and after various treatments, including sanitization and autoclaving procedures maintained its IgG binding properties, thus indicating a very high stability in terms of ligand leakage and degradation.

Adaptor Proteins, Signal Transducing↗

PubMed2002

Sequence-simplification and chimeric assembly: new models of peptide antigen modification.

Sequence-simplified variants of a 15-mer peptide antigen, identified by amino acid side chains in alternating positions were synthesized introducing glycine residues alternatively in the parent peptide sequence and used to induce antibodies in rabbit. They reacted to a significant extent with anti-parent peptide antibodies, and in addition, affinity purified antibodies against these halved forms recognized with similar affinity and specificity, the starting peptide in affinity chromatography, optical biosensor and enzyme linked immunosorbent assay (ELISA) experiments, while no cross-reactivity was detected between reduced antigens. These findings suggest that a peptide antigen can display two molecular surfaces of recognition, identified by side chains of residues in alternating positions. Each surface can even take part in antigen/antibody interaction independently, thus indicating the possibility to select and assembly sequence-simplified forms belonging to different epitopes, also deriving from different molecules, to generate new structures incorporating a two-fold antigen/antibody specificity. Two "chimeric" forms were then synthesized starting from the P15 and P13 complementary peptides, both able to bind interleukin 2. These structures, showing simultaneously trans-surfaces of recognition belonging to both parent forms, have been found to retain antigenic properties against antibodies of simplified P15 derivatives showing the same molecular surface of recognition. In addition, anti-chimeric antibodies recognized both P15 and P13 starting peptides, while no cross-antibody recognition was observed between chimeric antigens.

Amino Acid Sequence↗

PubMed2002

Novel molecular targets for systemic lupus erythematosus.

For a long time the complement cascade has been believed to be the predominant pathway to inflammation and tissue destruction in autoimmune diseases such as systemic lupus erythematosus. Recently, new evidences show that FcRs may share the primacy with complement cascade, playing an equal or greater role in the disease process. The generation of specific mouse strains deficient in individual components has clarified the different role played by complement and Fc receptors in their interaction with ICs, illustrating that complement is essential for innate immunity against microbial pathogens, requiring natural antibodies to mediate its protective effects, whereas FcyRs have evolved as the principal system for coupling antigen-antibody complexes to effector cells and initiate the inflammatory cascade. Validation of FcRs as new therapeutic targets for autoimmune diseases, in particular for Systemic Lupus Erythematosus (SLE), has been provided by a large number of studies where the biological action of soluble forms of FcyRs or of monoclonal antibodies targeting Fc receptors has been assessed. Additional support to the role of FcRs in SLE has been provided by data obtained with compounds derived from combinatorial chemistry, such as TG19320, a tetrameric tripeptide which interferes with IgG/FcgammaR interaction in vitro and prevents glomerulonephritis in vivo in a SLE susceptible mouse strain. These findings might open the way to new therapeutic approaches for disorders where the role of FcRs has been established, including not only autoimmune diseases like systemic lupus erythematosus, rheumatoid arthritis, multiple myeloma, but also acquired immunodeficiency syndrome (AIDS).

Animals↗