| Interleukin-6 which participates in multiple reactions in the body is a multifunctional cytokines,as well as an inflammation marker.It plays an important role in health and it is an important means of early diagnosis and screening of infectious diseases.Therefore,the measurement of the content of interleukin-6 in blood plays a significant guidance role in the supervision of several diseases,such as inflammatory responses,infectious diseases,tumor,etc.At present,there are multiple detection methods of interleukin-6,such as chemiluminescence immunoassay and ELISA which are the most common two with high detection sensitivity.The equipments of these methods are expensive,and they required professional operating staff.Fluorescent quantitation immunochromatography in this research is based on the quantitative determination method of immunology.This method is quick,delicate,peculiar,easy and low-cost and it has wide application prospects.In this study,interleukin-6 immune fluorescent latex was first prepared.Field emission scanning electron microscopy,dynamic light scattering and Zeta electric potential were used to discuss the stability of interleukin-6 immune fluorescent latex.The results showed when activator was EDC/NHS,active time 15 min,the reaction buffer pH value 5.5 MES,coupling time 2 h and the antibody concentration 0.6 mg/m L,the coupling ratio was the highest.The stability of immune fluorescent latex was enhance after it was coupled with IL-6 protein,studied by the data of field emission scanning electron microcopy,dynamic light scattering and Zeta potential tests.This study was carried out on the ELISA and fluorescence quantitative immunochromatography platform for a appropriate pair of antigen and antibody,eventually got a pair of coating antigen B1,antibody A2 and A3.The process optimum conditions of fluorescence quantitative immunochromatography stips were studied.Result showed optimum conditions were that antibody concentration of test line 0.5 mg/mL,spray volume of test line 2 ?L/cm,a75:100 ratio for adding proportion of sample and sample dilution,48 h for drying time of nitrocellulose membrane,a 1:10 ratio for adding proportion ofimmune fluorescent latex and working dilution,spray volume of conjugated pad 4 ?L/cm parallel jet line of 3 and 15 min for reation time.The standard curve showed a good linear with the R~2=0.99954,LOD and LOQ of the strip respectively were 8.494 pg/m L and 25.2789 pg/mL.The average recoveries were 96.51%?102.46%,103.06%,and the coefficients of variation respectively were 9.6%?8.57%?2.59%.The precision of intra-assay and inter-assay meet the requirements.The specificity and stability of the strip were good.In this study,the prepared kit was used for preliminary clinical application.The result showed the Kappa coefficient was 0.842,P<0.01.Two methods for the detection has good consistency,the correlation coefficient R~2=0.9652.The sensitivity,specific degree and the total coincidence of test kit's were 94.4%,97.6%,95% based on data of comparison of test kit and clinical standard method.The result also showed the correliation coefficient of serum/plasma,plasma/whole blood,serum/whole blood respectively were 1.2,0.61 and 0.732.Therefore,the preparation of fluorescence quantitative immunochromatographic kits are effective and can be used in point of care testing. |
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