Optimum conditions for in vitro production of interleukin 2 （IL 2） like activity from the Peripheral blood lymphocytes （PBL） of an Indian major carp, Labeo rohita were studied. Culture supernatants were generated ...Optimum conditions for in vitro production of interleukin 2 （IL 2） like activity from the Peripheral blood lymphocytes （PBL） of an Indian major carp, Labeo rohita were studied. Culture supernatants were generated by culturing the PBL in RPMI-1640 media supplemented with Glutamine and 10% fetal bovine serum （FBS） and stimulating with two different mitogens： concanavalin A （Con A） and Phytohaemagglutinin （PHA） at different concentrations separately. Significantly （P 〈 0.01） higher proliferation response was obtained from the culture supematant stimulated with concanavalin A （Con A） at a concentration of 10 lag mLl. The effect of phorbol myristate acetate （PMA） was also studied by co-stimulating PBL with Con A and PHA separately and it was found to synergistically enhancing the stimulation index with Con A whereas the stimulation index remain unchanged with PHA. The Con A （10 μg mLl） stimulated PBL were also cultured at different cell density, incubation period and incubation temperature in order to optimize the in vitro L. rohita IL2 production. The IL2 like activity was studied by lymphocyte proliferation assay on 72 h Con A blasts using WST based assay technique. Significantly （P 〈 0.01） higher stimulation indices were obtained when the PBL were cultured at a cell density of 1 × 10^6 cells mL^-1 for 30-36 h at an incubation temperature of 30 ℃. The IL2 like activity was purified by DEAE-Sepharose anion exchange chromatography and recorded between 70-130 mM NaCI with peak activity at 110 Mm NaCI. The molecular weight of the factor responsible for IL2 like activity was found to be 15-17 KD.展开更多
A quantitative method using fluorescence spectro-photometer to detect theinterleukin 2 reocptor （IL 2R） on the surface of mononuclear cells in normal humanperipheral blood is described. For expression of IL 2R, the ...A quantitative method using fluorescence spectro-photometer to detect theinterleukin 2 reocptor （IL 2R） on the surface of mononuclear cells in normal humanperipheral blood is described. For expression of IL 2R, the optimum conoentration of PHA andCon A were 100 μg/ml and 10 μg/ml respectively. The PHA induced effect on mononuclear cellswas better than Con A induced one. The peak value of IL 2R cxpression was at the 24th h afterstimulation The optimum dilution of the first antibody （anti-Tac） was 1: 50, while the dilution ofthe second antibody（fluorescein conjugated rabbit anti-mousc IgG） was 1: 32 Their optimumincubation period was 20 and 10 min repectively. The concentration of fluorcscein-conjugatedantibody per 1×10<sup>6</sup> mononuclear cells from the same normal human peripheral blood was5.22±0.45×10<sup>-10</sup> mol, and the coefficient of variation was 8. 6%. The concentration offluorescein-conjugated antibody per 1×10<sup>6</sup>PBMC from five healthy individuals was 4. 8±0. 97×10<sup>-10</sup> mol Therefore this assay system is stable and quite reproducible.展开更多
Background: Cytokines are mediators of diseases. Expression levels in the blood could be of clinical relevance. Objective: sIL-2Rα is used as a marker for different malignancies in human medicine. The aim of this stu...Background: Cytokines are mediators of diseases. Expression levels in the blood could be of clinical relevance. Objective: sIL-2Rα is used as a marker for different malignancies in human medicine. The aim of this study was to show if sIL-2Rα is detectable and if there is any correlation to different diseases in dogs. Methods: For this purposes sIL-2Rα concentrations in the blood were measured in healthy dogs, in dogs with different non-neoplastic diseases and benign tumors and in dogs with malignant tumors. Serum levels of sIL-2Rα were measured by using a human specific enzyme linked immunosorbent assay (ELISA). Results: Measurement of sIL-2Rα was successful in most of the samples. Dogs with diseases have significantly increased serum levels of sIL-2Rα compared to healthy controls. sIL-2Rα serum levels are higher in patients with non-neoplastic diseases and benign tumors than in those with malignant neoplasia. There is a strong correlation between sIL-2Rα and leukocyte count. Conclusion: Measurements of sIL-2Rα in serum may be helpful in detecting stages and grades of inflammation in the progression of disease. sIL-2Rα could actually not be used as an indicator for malignant diseases in dogs like in humans. The strong correlation between sIL-2Rα and the leukocyte count indicates the inflammatory response to the disease. This could be helpful in giving a prognosis in some cases, because the inflammatory reaction is of prognostic relevance in different diseases including malignant and non-malignant neoplasia. Although the results of our research studies were very promising, further studies should be performed with a canine ELISA.展开更多
文摘Optimum conditions for in vitro production of interleukin 2 （IL 2） like activity from the Peripheral blood lymphocytes （PBL） of an Indian major carp, Labeo rohita were studied. Culture supernatants were generated by culturing the PBL in RPMI-1640 media supplemented with Glutamine and 10% fetal bovine serum （FBS） and stimulating with two different mitogens： concanavalin A （Con A） and Phytohaemagglutinin （PHA） at different concentrations separately. Significantly （P 〈 0.01） higher proliferation response was obtained from the culture supematant stimulated with concanavalin A （Con A） at a concentration of 10 lag mLl. The effect of phorbol myristate acetate （PMA） was also studied by co-stimulating PBL with Con A and PHA separately and it was found to synergistically enhancing the stimulation index with Con A whereas the stimulation index remain unchanged with PHA. The Con A （10 μg mLl） stimulated PBL were also cultured at different cell density, incubation period and incubation temperature in order to optimize the in vitro L. rohita IL2 production. The IL2 like activity was studied by lymphocyte proliferation assay on 72 h Con A blasts using WST based assay technique. Significantly （P 〈 0.01） higher stimulation indices were obtained when the PBL were cultured at a cell density of 1 × 10^6 cells mL^-1 for 30-36 h at an incubation temperature of 30 ℃. The IL2 like activity was purified by DEAE-Sepharose anion exchange chromatography and recorded between 70-130 mM NaCI with peak activity at 110 Mm NaCI. The molecular weight of the factor responsible for IL2 like activity was found to be 15-17 KD.
文摘A quantitative method using fluorescence spectro-photometer to detect theinterleukin 2 reocptor （IL 2R） on the surface of mononuclear cells in normal humanperipheral blood is described. For expression of IL 2R, the optimum conoentration of PHA andCon A were 100 μg/ml and 10 μg/ml respectively. The PHA induced effect on mononuclear cellswas better than Con A induced one. The peak value of IL 2R cxpression was at the 24th h afterstimulation The optimum dilution of the first antibody （anti-Tac） was 1: 50, while the dilution ofthe second antibody（fluorescein conjugated rabbit anti-mousc IgG） was 1: 32 Their optimumincubation period was 20 and 10 min repectively. The concentration of fluorcscein-conjugatedantibody per 1×10<sup>6</sup> mononuclear cells from the same normal human peripheral blood was5.22±0.45×10<sup>-10</sup> mol, and the coefficient of variation was 8. 6%. The concentration offluorescein-conjugated antibody per 1×10<sup>6</sup>PBMC from five healthy individuals was 4. 8±0. 97×10<sup>-10</sup> mol Therefore this assay system is stable and quite reproducible.
文摘Background: Cytokines are mediators of diseases. Expression levels in the blood could be of clinical relevance. Objective: sIL-2Rα is used as a marker for different malignancies in human medicine. The aim of this study was to show if sIL-2Rα is detectable and if there is any correlation to different diseases in dogs. Methods: For this purposes sIL-2Rα concentrations in the blood were measured in healthy dogs, in dogs with different non-neoplastic diseases and benign tumors and in dogs with malignant tumors. Serum levels of sIL-2Rα were measured by using a human specific enzyme linked immunosorbent assay (ELISA). Results: Measurement of sIL-2Rα was successful in most of the samples. Dogs with diseases have significantly increased serum levels of sIL-2Rα compared to healthy controls. sIL-2Rα serum levels are higher in patients with non-neoplastic diseases and benign tumors than in those with malignant neoplasia. There is a strong correlation between sIL-2Rα and leukocyte count. Conclusion: Measurements of sIL-2Rα in serum may be helpful in detecting stages and grades of inflammation in the progression of disease. sIL-2Rα could actually not be used as an indicator for malignant diseases in dogs like in humans. The strong correlation between sIL-2Rα and the leukocyte count indicates the inflammatory response to the disease. This could be helpful in giving a prognosis in some cases, because the inflammatory reaction is of prognostic relevance in different diseases including malignant and non-malignant neoplasia. Although the results of our research studies were very promising, further studies should be performed with a canine ELISA.
