ABSTRACT
Six unique phage antibodies to human TNF have been selected from a combinatorial library of human single chain fragment variable. ELISA and Western-blotting was used to study selected phage antibodies binding with TNF. The specificity of selected antibodies was determined by binding with interferon alpha and gamma, bovine serum albumin, ovalbumin and ubiquitin. Two antibodies, sA1 and sB3, were converted into a soluble single-chain antibody form and their affinity was 2.5 and 13.7 nM respectively.
Subject(s)
Single-Chain Antibodies/immunology , Tumor Necrosis Factors/immunology , Amino Acid Sequence , Antibody Affinity/immunology , Antibody Specificity/immunology , Cytokines/immunology , Humans , Molecular Sequence Data , Ovalbumin/immunology , Peptide Library , Serum Albumin, Bovine/immunology , Single-Chain Antibodies/genetics , Single-Chain Antibodies/isolation & purification , Ubiquitin/immunologyABSTRACT
Twenty unique phage antibodies to human tumor necrosis factor alpha were selected from a naive combinatorial library of human single chain fragment variable. Analysis of gene segments encoding selected antibodies shown that repertoire of variable domains of heavy and light chains included variable domains of both naive autoantibodies and antibodies produced as a result of somatic hypermutagenesis.
Subject(s)
Antibodies, Monoclonal/genetics , Autoantibodies/genetics , Bacteriophage M13 , Gene Library , Immunoglobulin Variable Region/genetics , Single-Chain Antibodies/genetics , Tumor Necrosis Factor-alpha , Antibodies, Monoclonal/immunology , Autoantibodies/immunology , Humans , Immunoglobulin Variable Region/immunology , Single-Chain Antibodies/immunology , Somatic Hypermutation, Immunoglobulin/physiologyABSTRACT
Four unique phage single-chain variable fragments (scFvs) to recombinant human interleukin 18 (IL-18) has been selected from a naïve combinatorial library of human scFvs. Binding of unique phage antibodies with IL-18 was tested by ELISA and Western-blotting. No cross reactivity with tumor necrosis factor α, interferons α and γ was shown for the selected antibodies. The gene segments encoding V(D)J regions of selected antibodies exhibited a high degree of homology to germline genes, therefore we suggest that the selected scFv's belong to repertoire of naïve autoantibodies.
Subject(s)
Autoantibodies/immunology , Interleukin-18/immunology , Peptide Library , Recombinant Proteins/immunology , Single-Chain Antibodies/immunology , Amino Acid Sequence , Autoantibodies/genetics , Blotting, Western , Enzyme-Linked Immunosorbent Assay , Humans , Immunoglobulin Heavy Chains/chemistry , Immunoglobulin Heavy Chains/genetics , Immunoglobulin Heavy Chains/immunology , Immunoglobulin Light Chains/chemistry , Immunoglobulin Light Chains/genetics , Immunoglobulin Light Chains/immunology , Interleukin-18/genetics , Molecular Sequence Data , Polymorphism, Restriction Fragment Length , Recombinant Proteins/genetics , Single-Chain Antibodies/geneticsABSTRACT
KatG Ser3 15Thr mutation is one of the main reasons of resistance to isoniazid in Mycobacterium tuberculosis isolates. The frequency rate of the above mutation among isoniazid-resistant isolates made 94% in Novosibirsk Region and 93% in Kemerovo Region. The use of PCR-restriction fragment length polymorphism (PCR-RFLP) can be regarded as an adequate method for rapid screening of isoniazid resistance in Mycobacterium tuberculosis isolated in the West-Siberian Region.
