Compartmentalized Linker Array: A Scalable and Transferrable Microarray Format for Multiplexed Immunoassays

Microarrayshave been widely used for multiplexed bioassays. Fabricationof a conventional microarray typically requires a complex microarrayspotter, using which nanoliter bioreagent (e.g., antibody and cells)droplets are delivered onto a glass slide. However, arraying a delicatebioreagent in nanoliter volumes could cause the loss of bioactivityand needs a complex microarray spotter. Further, mixing of differentbioreagents in a multiplexed assay leads to cross-reactions, producingfalse positive signals that impair assay reproducibility and scalability.In this work, we propose a new microarray format, named compartmentalizedlinker array (CLA), that consists of pre-prepared storablemicroarrays of chemical linkers in microliter compartments. CLA canbe used for binding and patterning bioreagents into microarrays bysimply pipetting and incubating bioreagent solutions in compartments.Using commonly used aminosilane linker-based antibody microarray,we developed CLA and demonstrated its application for a multiplexedsandwich immunoassay measuring three cancer-related proteins. A two-phaseblocking system was established for de-activating background regionson glass where no linker molecules are present. Storage conditionsof the CLA chip were explored and demonstrated for long-term storage.In a multiplexed immunoassay, low pg/mL sensitivity was achieved forall the three proteins, comparable to those of conventional assays.Moreover, CLA can be potentially used for other applications beyondprotein assays, making microarray technology transferrable and widelyavailable for the biological and biomedical research community.

Automated summary

A new microarray format, named compartmentalized linker array (CLA), was proposed, which consists of pre-prepared microarrays of chemical linkers in microliter compartments. CLA can bind and pattern bioreagents into microarrays by simply pipetting and incubating bioreagent solutions in compartments. The application of CLA was demonstrated for a multiplexed sandwich immunoassay measuring three cancer-related proteins.