1.Effect of different isolations for nucleic acid and protein complex on affinity of enriched library in SELEX experiment
Chengxiang HU ; Changguo GU ; Xudong ZHU ; Lei LI ;
Journal of Third Military Medical University 2003;0(08):-
Objective To compare the effect of different partition of nucleic acid protein complex on the affinity of enriched library in systematic evolution of ligands by exponential enrichment (SELEX). Methods Two protocols were adopted to select the enriched library to transforming growth factor ? receptor Ⅱ(TGF ? RⅡ). Protocol 1: protein was absorbed on the surface of 96 well plate; then, selection was carried out; the binding nucleotide acids were eluted from the supporter directly. Protocol 2: selection was done in solution; nucleotide acid protein complex was captured in nitrocellulose membrane; the binding nucleotide acids were obtained from membrane. Filter biding assay and gel shift assay were performed to detect the affinity of the enriched ssDNA library from different protocols. Results After 4 rounds of selection, the affinity to TGF ? RⅡ was obviously improved in the enriched library from protocol 2 compared with the initial library, while no such improvement was found in the enriched library from protocol 1. Conclusion In the SELEX experiment, the way of selection in solution, then partition of the binding nucleotide acids in filter is easier to enrich the binding fragment from initial ssDNA random library, compared with the way of fixation of target protein to solid supporter, then selection between the solid phase and liquid phase and elution of the binding nucleotide acids from supporter.
2.Effect of repeated measles vaccine injection on interleukin-12, interleukin-13 levels in asthmatic children
Weiping TAN ; Shaoliang HUANG ; Xiandi MAI ; Li ZHAO ; Baoqing WU ; Huarong HUANG ; Changguo ZHU ; Yanfeng WU
Chinese Journal of Pathophysiology 1986;0(02):-
AIM and METHODS: To study the immunological effect of measles vaccine therapy on asthmatic children, we examined the changes of interleukin-12 , interleukin-13 and total serum IgE levels in plasma and cultured peripheral blood mononuclear cells(PBMC) supernatant by means of ELISA in 13 mild-moderate asthmatic children treated with measles vaccine. Results were compared with 12 anti-symptomatic treatment mild-moderate asthmatic children and 17 normal children control group. RESULTS:After measles vaccine treatment, IL-13 and total serum IgE levels decreased remarkably, statistically lower than that of group receiving only anti-symptomatic treatment. There was no statistical difference in IL-12 level between the two group. Correlation analysis: 1)IL-12 level of plasma was negatively correlated to the level of serum total IgE, there was no correlation of supernatant IL-12 in PBMC to the total serum IgE; 2)IL-13 levels in plasma and PBMC were positively correlated to the level of total serum IgE; 3) IL-12 level was negatively correlated to IL-13. CONCLUSION: Measles vaccine could down-regulate IL-13 level, therefore decrease total IgE synthesis, but not affect IL-12 level in asthmatic children.
3.Preliminary study on levels of IL-4,IL-12 and IgE in children with bronchial asthma
Li ZHAO ; Xiandi MAI ; Weiping TAN ; Huarong HUANG ; Shaoliang HUANG ; Changguo ZHU
Journal of Clinical Pediatrics 2001;(1):9-10,40
In this study,IL-4 and IL-12 levels in plasma and peripheral blood mononuclear cells(PBMC) as well as serum IgE level were prospectively assessed with double-antibody sandwich ELISA technique in children with asthma during the attack group (30 cases)and the interval group(12 cases).The results observed revealed that serum IgE level and IL-4 level in plasma and PBMC after PHA and LPS provocation during both of the attack stage and the interval stage were found to be evidently higher than those of the normal control group (P<0.01).Otherwise,IL-12 level during both stages was much lower than that of the normal control(P<0.01).In other hand,there were found to have a significant difference in all these 3 data between attack and interval stages.Thus, the conclusion indicates that there might be an imbalance of IL-4,IL-12 and IgE level in children with asthma during both of the attack and the interval stages.