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J Vis Exp. 2013 Nov 23;(81):e50935. doi: 10.3791/50935.

High-throughput flow cytometry cell-based assay to detect antibodies to N-methyl-D-aspartate receptor or dopamine-2 receptor in human serum.

Author information

1
Institute for Neuroscience and Muscle Research, The Kids Research Institute at the Children's Hospital at Westmead, The University of Sydney.

Abstract

Over the recent years, antibodies against surface and conformational proteins involved in neurotransmission have been detected in autoimmune CNS diseases in children and adults. These antibodies have been used to guide diagnosis and treatment. Cell-based assays have improved the detection of antibodies in patient serum. They are based on the surface expression of brain antigens on eukaryotic cells, which are then incubated with diluted patient sera followed by fluorochrome-conjugated secondary antibodies. After washing, secondary antibody binding is then analyzed by flow cytometry. Our group has developed a high-throughput flow cytometry live cell-based assay to reliably detect antibodies against specific neurotransmitter receptors. This flow cytometry method is straight forward, quantitative, efficient, and the use of a high-throughput sampler system allows for large patient cohorts to be easily assayed in a short space of time. Additionally, this cell-based assay can be easily adapted to detect antibodies to many different antigenic targets, both from the central nervous system and periphery. Discovering additional novel antibody biomarkers will enable prompt and accurate diagnosis and improve treatment of immune-mediated disorders.

PMID:
24300941
PMCID:
PMC3992012
DOI:
10.3791/50935
[Indexed for MEDLINE]
Free PMC Article

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