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Objective@#To establish a quantitative assay of serum Golgi protein 73 (GP73) using xMAP technology and evaluate its performance.@*Methods@#Monoclonal antibodies against GP73 were prepared and purified, and antibody pair screening was performed by double-antibody sandwich enzyme-linked immunosorbent assay. The screened antibodies were used to construct a Luminex liquid chip detection system, and the analysis performance of the detection system was evaluated. The serum levels of GP73 were detected in 90 clinical samples from healthy controls and patients with chronic hepatitis B infection (CHB) and hepatocellular carcinoma (HCC).@*Results@#Five anti-GP73 monoclonal antibodies were prepared and purified, and 5 antibody pairs were successfully screened. The Luminex liquid chip detection system of GP73 was successfully constructed using 8F10D1 and 10B9F11 antibody pairs. The analytical performance evaluation showed that the sensitivity of this system was 0.25 ng/ml and the dynamic range was 0.25-100 ng/ml. No cross reactivity was observed. The intra- and inter-assay variation for GP73 was <8% and <11%, respectively. The recovery was 83%-92%. The linear regression equation was y=1.141x+ 6.436 (r2=0.998 4, P<0.001). The GP73 concentrations in the serum samples of healthy control, CHB group, and HCC group were 42.8 (38.68, 55.90) ng/ml, 61.49 (43.59, 81) ng/ml, and 122.78 (49.36 liter, 264.55) ng/ml, respectively. The levels of GP73 in HCC group were significantly higher than those in CHB group and healthy controls (P<0.05). Moreover, the levels of GP73 in CHB group were significantly higher than those in healthy controls (P<0.05).@*Conclusions@#A liquid chip detection system of GP73 was successfully constructed. It provides a powerful tool for the clinical application of GP73 in the future.
ABSTRACT
ObjectiveTo study the dynamic express of CD57 on T cell of PBMC and clinical significance in acute HIV infection.MethodsSeventeen patients with acute HIV infection were enrolled study randomly diagnosed from 2006.11 to 2009.12 and 15 healthy donors as control group.The PBMCs from 1th,3th and 6th during acute infection were collected.The proportion of CD3+CD57+T lymphocytes,CD3+ CD8+CD57 +T lymphocytes and CD3 + CD4 + CD57 + T lymphocytes were evaluated by flow cytometric analysis with three or double color staining.The relationship between the proportion of CD57+ T phenotypes and virus load and CD4+T cells count was analyzed.ResultsThe proportion of CD57+T lymphocytes in PMBC in 1th,3th and 6th during acute HIV acute was 15.24％ ±1.49％,13.51％ ±2.45％ and 14.65％ ±1.83％,respectively,and was higher than normal control group and the difference was significantly(P＜0.0001 ).The proportion of CD8+ CD57+T lymphocytes was 7.79％ ±2.10％ and 9.88％ ±2.36％ in 1th and 3th month during acute infection,respectively.The proportion of CD8+ CD57+T lymphocytes in 1th and 3th month during acute infection were positive relationship with virus load in corresponding time,and R2 was 0.3700 and 0.3768,and P value was 0.0096 and 0.0088,respectively.The proportion of CD8+CD57+T lymphocytes in the 1th and 3th month during acute infection was negative relationship with CD4+T lymphocytes count.The R2 was 0.3768 and 0.4235,and P value was 0.0215 and 0.0017,respectively.In 6 rapid progressors and 11 no rapid progressors on the 1th month after HIV infection,CD8+ CD57+T lymphocytes percentage was 11.20％±2.21％ and 6.16％ ±1.09％,respectively,and CD4+CD57+T lymphocytes percentage 2.79％ ±0.31％and 1.40％ ±0.30％,respectively.Both CD8+CD57+T and CD4+CD57+T in rapid progressors were higher than no rapid progressors,and P value was 0.0338 and 0.0106,respectively.ConclusionCD57+ T lymphocytes percentage in peripheral blood increase in acute HIV infection patients,in which the increasing CD8+CD57+T lymphocyte may mirror the dynamic of HIV replication and CD4+T cell count.The CD57 high express on T lymphocyte on the early HIV acute infection predicts rapid progression.