In some cases, proper fluorescent and reporter intensity measurements are compromised in the presence of a large concentration of floating (unbound) labeled species in the incubation solution, whose signal can overwhelm the target-specific signal from the captured targets. In conventional fluorescent-based microarrays and other extrinsic reporter-based (label-based) biosensors assays, the detection of captured analytes is usually carried out after the incubation step. Presently, densely packed biosensor arrays which detect thousands of different analytes simultaneously (microarrays) may be popular in Genomics, Proteomics, molecular diagnostics, and systems biology. Biosensors in parallel may compensate for limited detection performance. An advantage of biosensors is their capability to be implemented in parallel and in an array format. The performance of biosensors in terms of signal to noise ratio and dynamic range is generally constrained by the characteristics of the molecular recognition layer which captures the target analytes, and not by the transducer and read-circuitry. Affinity-based biosensors exploit selective binding and interaction of certain bio-molecules (recognition probes) to detect specific target analytes in biological samples. It is of great interest to efficiently multiplex the amplification process and thus allow for multiple target amplification and quantification.īiosensor detection systems take advantage of the selective interaction and binding (affinity) of certain biological molecules to identify molecular structures and furthermore measure levels of different analytes such as toxins, polymers, hormones, DNA strands, proteins, and bacteria. These assays have become a powerful tool in molecular biology and genomics, since they can increase the number of copies of target molecules with great specificity. Nucleic acid target amplification assays such as polymerase chain reaction process (PCR), in principle, amplify and replicate specific sequences of nucleic acids of a DNA template in vitro. 5, 2006, the disclosures of which are incorporated herein by reference in their entireties. 9,133,504, which claims the benefit of U.S. 10, 2015, which is a continuation of U.S. 24, 2006 and is a continuation-in-part of U.S. 8,637,436, which claims the benefit of U.S. 10,106,839, which is a divisional of U.S. 29, 2017, which is a continuation-in-part of U.S. 10, 2019, which is a continuation-in-part of U.S. This application is a continuation of U.S. The present disclosure also provides a fully integrated bioarray for detecting real-time characteristics of affinity based assays. Also provided herein are biosensor arrays, systems and methods for affinity based assays that are able to simultaneously obtain high quality measurements of the binding characteristics of multiple analytes, and that are able to determine the amounts of those analytes in solution. The measured amount of amplicon produced can be used to determine the original amount of the nucleic acid sequences in the sample. The amount of amplicon corresponding to the multiple nucleic acid sequences can be measured in real-time during or after each cycle using a real-time microarray. The nucleic acid sequences in the sample are simultaneously amplified, for example, using polymerase chain reaction (PCR) in the presence of an array of nucleic acid probes. This present disclosure provides methods and systems for measuring the concentration of multiple nucleic acid sequences in a sample.
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