This article has not been peer reviewed.Deposited research article
Protein microarrays for highly parallel detection and quantitation of specific proteins and antibodies in complex solutions
1 Department of Biochemistry, Stanford University School of Medicine, Stanford, CA 94305, USA
2 Department of Genetics, Stanford University School of Medicine, Stanford, CA 94305, USA
3 Howard Hughes Medical Institute
4 Current address: The Van Andel Research Institute, 333 Bostwick, NE, Grand Rapids, Mi 49503, USA
Genome Biology 2000, 1:preprint0001-preprint0001.22 doi:10.1186/gb-2000-1-6-preprint0001
This was the first version of this article to be made available publicly. A peer-reviewed and modified version is now available in full at http://genomebiology.com/2001/2/2/research/0004Published: 17 November 2000
We describe a method for printing protein microarrays, and using these microarrays in a comparative fluorescence assay to measure the abundance of many specific proteins in complex solutions. A robotic device was used to print hundreds of specific antibody or antigen solutions in an array on the surface of derivatized microscope slides. Two complex protein samples, one serving as a standard for comparative quantitation, and the other representing an experimental sample in which the concentrations of specific proteins were to be measured, were labeled by covalent attachment of spectrally-resolvable fluorescent dyes. Specific antibody-antigen interactions localized specific components of the complex mixtures to defined cognate spots in the array, where the relative intensity of the fluorescent signals representing the experimental sample and the reference standard provided a measure of each protein's abundance in the experimental sample. To characterize the specificity, sensitivity and accuracy of this assay, we analyzed the performance of 115 antibody/antigen pairs.
50% of the arrayed antigens, and 20% of the arrayed antibodies, provided specific and accurate measurements of their cognate ligands at or below concentrations of 1.6 µg/ml and 0.34 µg/ml, respectively. Some of the antibody/antigen pairs allowed detection of the cognate ligands at absolute concentrations below 1 ng/ml, and partial concentrations of less than 1 part in 106, sensitivities sufficient for measurement of many clinically important proteins in patient blood samples.
Protein microarrays can provide a simple and practical means to characterize patterns of variation in hundreds or thousands of different proteins, in clinical or research applications.