| Chronic renal diseases (CRDs) always involve in immunological factors and inflammation medium and non-immunological factors as well. Cytokine plays a very important role during the occurance and development of these diseases. Hence, We detect the serum neoptin level which represents T cell-mediated immune(CMI), the serum interleukin-10 (IL-lO)level which represents B CMI and the serum level of transforming growth factor P i(TGF P i) whose level in renal tissue correlated with fibrosis degree in patients with CRD and discuss their role and significance . OBJECTIVETo detect the serum levels of NP, IL-10 and discuss their role and significance in patients with CRD. MATERIAL AND METHOD1. Investigated objects and divided into groups: every patient is that of an inpatient with CRD. According to the ratio of 24 hours urine protein, the levels of serum albumin and the serum creatinine and/or the ratio of clean creatinine, all the patients in current were divided into 3 groups: nephritic syndrome (NS),non-nephritic syndrome (non-NS) and chronic renal insufficiency (CRI). Meanwhile, to compare their varieties of the detected markers in patients with various pathologic diagnoses, we divided the 23 cases, which have pathologic diagnosis into 4 categories: proliferative non-proliferative, immune complex (1C) mediated and non-IC-mediated. The proliferative groups consisted of immunoglobulin A (Ig A) nephropathy, membranoproUferative GN (MPGN), mesangial proliferative glomerulonephritis (MsPGN). The non-proliferative groups included minimal change disease (MCD), focal segmental glomerulosclerosis (FSGS) and membranous nephropathy(MN). The immune complex (1C) mediated groups included MM, IgA nephropathy, MPGN. The non-IC-mediated consisted of MCD, FSGS, mesangial proliferative glomerulonephritis(GN)2. Sample collection: the whole blood 3-5ml was taken at the early morning in the fasting state. The serum separated was cold storied to take the test. The control samples came from duty blood donors. The serum separated was cold storied to take the test.3. Test of the serum levels of IL-10, TGF Pi by the mean of the double antibody Enzyme-linked immunoadsordent assay: Testing IL-10, taking out the wells,we pipette lOOul of sample, control or standard in every well. The wells were covered with a foil and incubated for 120 minutes at room temperature, and then washed four times with wash buffer. Pipette lOOul of biotinylation antibody solute in every well and incubate them for 120 minutes at room temperature and then wash them four times with wash buffer. Pipette lOOul of enzyme conjugate in every well and incubate them for 30 minutes at room temperature and wash them four times with wash buffer. Pipette 50ul of A substrate solution and B substrate solution in every well and incubate them for 10 minutes at roomtemperature, avoiding light. Wash them four times with wash buffer. Pipette 50ul of the stop solution hi every well. Mix the solutions briefly and read their absorbance. The real absorbance is the balance after the tested absorbance subtracted that of the empty well. According to the absorbance of the standard and their normal concentration, make a standard curved line. According to the absorbance, make out the concentration of IL-10. When we test the concentration of TGF P i,we should pre-treat(dilution 1:50) the samples.4. Test the serum NP level by the mean of competitive inhibition. Take out the wells, pipette lOul of sample, control or standard hi every well; and then pipette lOOulof Enzyme conjugate (1:20), and pipette SOulof NP antiserum one by one in every well. The wells were Covered with a black foil and incubated for 90 minutes at room temperature on an orbital shaker. Decant the supernatant^ washed them 3 times by wash buffer. Pipette 200ul of TMB substrate solution hi every well and incubate them for 10 minutes at room temperature. Pipette 200ul of TMB stop solution in every well, mix them briefly and read their absorbance. According to the absorbance, make out the concentration of NP.
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