Biochip point-of-care device for sepsis diagnostics

We report about a point-of-care device for the diagnosis of sepsis. For this a useful set of diagnostic parameters must be analyzed in parallel, to combine the diagnostic benefit of each parameter. By this the severity of sepsis can be diagnosed, predictive indicators can be gathered and it is possible to differentiate between viral and bacterial sepsis being all essential information for septic patients. Therefore the protein biomarkers C-reactive protein (CRP), interleukin 6 (IL-6), procalcitonin (PCT) and neopterin (NPT) are integrated in a multiparameter on-chip immunofluorescence assay. Furthermore important criteria for point-of-care testing (POCT) such as low testing time, small sample volume, compactness, cost efficiency, accurate fluid handling, automated operation, sensitive detection and good reproducibility are closely examined. The measurement system is based on Total Internal Reflection Fluorescence (TIRF) by use of special biochips. The system reads out microarray based multiparameter immunofluorescence assays. To characterize the device several standard curves were generated by three-fold replicates in a serum model with 4% human serum albumin and human plasma containing spiked standard analytes. PCT and IL-6 are carried out in a sandwich assay format whereas NPT and CRP are processed in a binding inhibition format. Both assay types are processed in parallel by the fluidic unit. The assay time for a single multiparameter assay is 25 min. An accurate execution of the two assays formats in parallel needs several fluid handling steps like dilution, mixing, metering, separating, pre-incubating and incubating to be carried out. The protein chip is processed fully automated using 10-75 ml human plasma or serum sample. An additional important demand for POCT is the accuracy and precision of the measurements. This paper shows a method how imprecision and inaccuracy can be drastically reduced by using on-chip reference assay signals to reference the sepsis parameter signals. The on-chip reference assay runs parallel to the multiparameter assay. The method is analyzed with standard curves using human plasma matrix. To investigate the performance of a new a point-of-care device, it is necessary to run the assays with clinical specimen of healthy and pathological patients. For this the IL-6 levels were first investigated by established clinical systems in two different hospitals and then compared with the new platform. It was possible to resolve the clinical relevant IL-6 levels and a required LOD in the lower pg/ml range was achieved.