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Instrument-free protein microarray fabrication for accurate affinity measurements -preprint.pdf (957.47 kB)
Instrument-Free Protein Microarray Fabrication for Accurate Affinity Measurements
Preprints are manuscripts made publicly available before they have been submitted for formal peer review and publication. They might contain new research findings or data. Preprints can be a draft or final version of an author's research but must not have been accepted for publication at the time of submission.
Protein microarrays have gained popularity as an attractive tool for
various fields, including drug and biomarker development, and diagnostics.
Thus, multiplexed binding affinity measurements in microarray format has become
crucial. The preparation of microarray-based protein assays relies on precise dispensing
of probe solutions to achieve efficient immobilization onto an active
surface. The prohibitively high cost of
equipment and the need for trained personnel to operate high complexity robotic
spotters for microarray fabrication are significant detriments for researchers,
especially for small laboratories with limited resources. Here, we present a
low-cost, instrument-free dispensing technique by which users who are
familiar with micropipetting can manually create multiplexed protein assays
that show improved capture efficiency and noise level in comparison to that of
the robotically spotted assays. In this study, we compare the efficiency of
manually and robotically dispensed α-Lactalbumin probe spots by analyzing
the binding kinetics obtained from the interaction with anti-α-Lactalbumin
antibodies, using the interferometric reflectance imaging sensor platform. We
show that the protein arrays prepared by micropipette manual spotting meet and
exceed the performance of those prepared by state-of-the-art robotic spotters.
These instrument-free protein assays have higher binding signal (~4-fold
improvement) and a ~3-fold better signal-to-noise ratio (SNR) in binding curves,
when compared to the data acquired by averaging of 75 robotic spots
corresponding to the same effective sensor surface area. We demonstrate the
potential of determining antigen-antibody binding coefficients in 24-multiplexed
chip format with less than 5% measurement error.
The authors declare the following competing financial interest(s): Prof. M. Selim Ünlü is the principal investigator of the technology translation grants listed in the funding information. He is the founder of a startup company (iRiS Kinetics, Inc.) for the commercialization of the IRIS multiplexed affinity technique.