Multiplexed ELISA

What are Multiplexed ELISAs?

Enzyme Linked Immunosorbant Assays (ELISA) have been used for decades in research and development, disease diagnosis and other critical fields in sciences.

ELISAs are simply a binding event between an antibody and an antigen with an enzymatic detection method. They can be used to detect proteins, peptides, lipids and any antigen that can be recognized by an antibody.

Quansys Multiplexed ELISAs are based on tried and proven sandwich (capture) ELISA techniques. This process uses two antibodies. One antibody is first bound to a solid phase (i.e. micro-well plate). The sample is incubated on the surface, allowing the target analyte and bound antibody to associate. Following this, each well is incubated with a second enzyme-linked antibody. The second antibody binds to a different region of the target analyte. This reaction is detected by monitoring the activity of the enzyme, which is proportional to the amount of target analyte present in the sample.

Conventional ELISAs, though allowing the researcher to test many samples at one time, can test only one type of analyte in a single test. This makes testing multiple analytes costly with respect to time, reagents, and sample volume. These factors and the importance of profiles when working with multiple analytes have prompted many researchers to develop assays in multiplexed formats. These formats have included fluorescent bead-based assays, micro titer plate arrays and slide-based protein arrays. These methodologies, in contrast to the conventional ELISA, allow the researcher to test many analytes at one time, but test only one or a few samples at a time, reducing the throughput of the assay.

The Quansys ME™ (Multiplexed ELISA) Array is a fully quantitative ELISA-based test where up to 25 distinct capture antibodies have been absorbed to each well of a 96-well plate in a defined array. This array is composed of 20 nanoliter spots with 350µm diameters and a pitch of 650µm between spots. Each spot represents a different distinct capture antibody population. Using less than 30 µl of sample, up to 84 different samples can be assayed for all 25 unique analytes in less than 2.5 hours. Sensitivity is system dependent. It typically ranges between 30 pg/ml to less than 1 pg/ml. All of the antibodies used in the ME™ arrays have been subject to a rigorous and comprehensive cross reactivity protocol and verified to be non-cross reactive with any other system on the array. Detection of this array is performed using a CCD based gel documentation system or the Quansys Imaging System. The image is then processed using Quansys Q-View™ Software and concentrations for each analyte are output for the sample.