Production, characterization, and applications of two novel monoclonal antibodies against human interleukin-28A.
Interleukin-28A (IL-28A) is a novel cytokine discovered in recent years and has been shown to have antiviral activity. In this study, IL-28A complementary DNA was inserted into prokaryotic expression vector pET-44 Ek/LIC. The Nus-S-His-tagged IL-28A fusion protein was expressed in Escherichia coli BL21 (DE3) in the soluble fraction. The fusion protein was purified by S-protein agarose affinity chromatography, and the fusion tag was removed from recombinant IL-28A by cleavage with thrombin. To prepare specific monoclonal antibody against human IL-28A, BALB/c mice were immunized with IL-28A, and hybridoma cell lines were obtained by fusing mouse spleen cells with myeloma NS-1 cells. Two strains of hybridoma cells, which produced the anti-human IL-28A antibodies 1B9 and 4B5 were obtained. They are IgM isotype and working in western blot analysis and enzyme-linked immunosorbent assay. In the present study, it was shown for the first time that human umbilical vein endothelial cells treated with interferon-alpha and poly(I:C) express IL-28A protein assessed by flow cytometry and immunofluorescent staining techniques. Immunohistochemistry showed that macrophage-like cells in colon and lung tissue and alveolar epithelial cells in lung tissue contain IL-28A, indicating a novel mechanism for both cell types to carry out their antivirus or antitumor functions.
Li
M
M
Allergy and Inflammation Research Institute, Shantou University Medical College, Shantou, People's Republic of China.
Chen
Y
Y
Huang
T
T
Liu
Y
Y
He
S
S
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000276:immunology*; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D002846:Chromatography, Affinity; D016207:Cytokines / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism; D005434:Flow Cytometry; D005822:Genetic Vectors; D006801:Humans; D007150:Immunohistochemistry; D007369:Interferon Inducers / Q000494:pharmacology; D016898:Interferon-alpha / Q000494:pharmacology; D007378:Interleukins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D011070:Poly I-C / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism; D000095022:Interferon Lambda
D000911:Antibodies, Monoclonal; D016207:Cytokines; D007369:Interferon Inducers; D016898:Interferon-alpha; D007378:Interleukins; D011070:Poly I-C; D011993:Recombinant Fusion Proteins; C000624353:interferon-lambda, human; D000095022:Interferon Lambda