Mass-Production and Characterization of Anti-CD20 Monoclonal Antibody in Peritoneum of Balb/c Mice.

PURPOSE: Monoclonal antibodies are important tools are used in basic research as well as, in diagnosis, imaging and treatment of immunodeficiency diseases, infections and cancers. The purpose of this study was to produce large scale of monoclonal antibody against CD20 in order to diagnostic application in leukemia and lymphomas disorders. METHODS: Hybridoma cells that produce monoclonal antibody against human CD20 were administered into the peritoneum of the Balb/c mice which have previously been primed with 0.5 ml Pristane. After twelve days, approximately 7 ml ascetic fluid was harvested from the peritoneum of each mouse. Evaluation of mAb titration was assessed by ELISA method. In the present study, we describe a protocol for large scale production of MAbs. RESULTS: We prepared monoclonal antibodies (mAbs) with high specificity and sensitivity against human CD20 by hybridoma method and characterized them by ELISA. The subclass of antibody was IgG2a and its light chain was kappa. Ascetic fluid was purified by Protein-A Sepharose affinity chromatography and the purified monoclonal antibody was conjugated with FITC and Immunofluorescence was done for confirming the specific binding. CONCLUSION: The conjugated monoclonal antibody could have application in diagnosis B-cell lymphomas, hairy cell leukemia, B-cell chronic lymphocytic leukemia, and melanoma cancer stem cells.

Green tea (Camellia sinensis) administration induces expression of immune relevant genes and biochemical parameters in rainbow trout (Oncorhynchus mykiss).

Present study elucidates the efficacy of green tea (Camellia sinensis) on growth performance, immune and antioxidant systems and cytokine gene expression in rainbow trout tissues. Green tea was supplemented at 20, 100, and 500 mg kg(-1) diet and fed to fish (average weight: 23.5 g) for 35 days. No remarkable changes in growth performance were observed among all test groups. Lower lipid peroxidation product and higher superoxide dismutase activity were noted in fish received the medium dose of green tea. Significant increase in serum bactericidal activity and total protein were recorded in all treatment groups. All doses of green tea up-regulated Interleukin-1ß transcription in the spleen, while Interleukin-1ß mRNA level decreased significantly in the kidney of low dose of green tea. Interleukin-6 mRNA level was up-regulated in the spleen of high dose of green tea and liver of middle and high doses of green tea. High dose and medium dose of green tea up-regulated the interleukin-8 transcription in the kidney and liver, respectively. Meanwhile, green tea inhibited the production of interleukin-10 in all treatment groups compared with control group. Medium dose of green tea up-regulated tumor necrosis factor-α transcription in all fish tissues, while high dose and low dose of green tea enhanced tumor necrosis factor-α mRNA levels in the kidney and spleen, respectively. Present study suggests that green tea especially at 100 mg kg(-1) feed may effectively enhance the antioxidant system and immune system in rainbow trout.

Seroepidemiology of toxoplasmosis in childbearing women of Northwest Iran.

Toxoplasma gondii causes the most common parasitic infection in the world. Congenital transmission, prenatal mortality and abortion are major problems of T. gondii. Prevalence of toxoplasmosis is high in Iran, especially in Azerbaijan. The current literature reviewed in this paper reveal results pertaining to various regions of Iran. The present cross-sectional e-study was designed to evaluate the seroprevalence of toxoplasmosis in childbearing women in Northwest Iran. We evaluated 1659 women in childbearing age from several cities in Northwestern Iran (Tabriz, Maragheh, Ahar, Marand, Sarab, Miane) from July 2009 to August 2010. Women aged between 20 and 40 years and seeking prenatal care were enrolled in the study. The subjects' sera were probed with indirect fluorescent antibody (IFA). A total of 1659 subjects were examined. Titres ranged from 1:100 to 1:800. In all, 899 subjects (54.13%) were seropositive. The highest frequency of seropositivity was shown in 1:200 dilution (36.08%) and in subjects from Maragheh (84% of 211 subjects). There was a direct linear relationship between seropositivity and age (p 0.001). Also, seroprevalence of toxoplasmosis was higher in subjects with primary school/lower educational level (p 0.001) and subjects living in rural regions (p 0.001). Overall, more than 50% of women in childbearing age were seropositive for toxoplasmosis in northwestern Iran. Increasing seroprevalence of toxoplasmosis with age was a predictable result due to longer exposure to the parasite. The relationship between increasing seroprevalence and lower educational level as well as living in rural areas is in line with the latest epidemiological findings, which also show such relationships due to lower socioeconomic status.

Increased IL-17A but decreased IL-27 serum levels in patients with multiple sclerosis.

BACKGROUND: Effector CD4+ T cell subsets play an important role in Multiple Sclerosis (MS). Interleukin-27 (IL-27) suppresses Th (Th1, Th2 and Th17) cells and dampens autoimmunity and tissue inflammation by promoting the generation of Type 1 regulatory T cells (Tr1). OBJECTIVE: To identify the relative levels of IL-27 and IL-17A in MS disease. METHOD: In a case-control study, venous blood was collected from forty MS patients and forty-three healthy subjects as control group. Serum levels of IL-27 and IL-17A were measured by ELISA method. RESULTS: A significant difference between serum IL-17A concentration in patients (120.68 ± 209.85 pg/ml) and control group (67.26 ± 117.76 pg/ml, p=0.016) was found. Serum IL-27 levels of the MS patients (159.7 ± 581.4 pg/ml) were significantly lower than control subjects (180.35 ± 507.84 pg/ml, p=0.001). CONCLUSION: Our findings show decreased levels of IL-27 against increasing IL-17A levels in patients group which may suggest a suppressive role of IL-27 on inflammatory process of MS.

Development and characterization of monoclonal antibodies against human CD20 in Balb/c mice.

BACKGROUND: Targeting CD20 antigen on B-lymphocytes provides good opportunity for management of the target cells in patients with B-cell malignancies. By the advent of hybridoma technology, monoclonal antibodies applications exert extensive changes in medical fields such as diagnosis, treatment and purification. OBJECTIVE: The prim aim of this study was to produce monoclonal antibody against CD20 for exploitation in diagnosis. METHODS: In this study, Balb/c mice were immunized with two peptides from extracellular domain of CD20. Poly Ethylene Glycol (PEG) fused spleen cells of the most immune mouse with SP2/0 (myeloma cells). Supernatant of hybridoma cells were screened for detection of antibody by ELISA. The desired clones were selected for limiting dilution (L.D). Afterward, specificity and cross reactivity of these antibodies were determined by immunological assay such as ELISA and western blot analysis (WB) and Immunofluorescence. Large scale of monoclonal antibodies was produced by ascetic fluid method. Monoclonal antibody was purified by chromatography then confirmed by SDS-PAGE. RESULTS: In this study, between five positive clone wells, 3 clones were chosen for limiting dilution. Limiting dilution product was one monoclone with absorbance about 2. CONCLUSIONS: These results indicate that such monoclonal antibodies against CD20 can be used in diagnosis of CD20 in the cells surface.