Bead-Based Immunoassay Developed to Measure Vimentin Autoantibodies

Vimentin is an intermediate filament protein generally expressed in the cytosol of many adult cell types, including leukocytes, fibroblasts and endothelial cells. Several tissue and/or injury-specific isoforms of vimentin are known to exist that may trigger autoimmune responses due to aberrant structural or conformational variations.

Scientists at the Rush University Medical Center (Chicago, IL, USA) evaluated the efficacy of the newly developed assay on discarded serum samples collected from 51 renal transplant patients with allograft dysfunction and compared the results with control specimens from 20 age and gender matched control individuals.

Magplex Microspheres (Luminex Corporation; Austin, TX, USA) were used in the development of this vimentin autoantibody assay and were processed in batches of 106 beads. Recombinant human vimentin (R&D Systems, Minneapolis, MN, USA) were added to the MagPlex beads and serum samples were processed. Assay performance and microsphere validation was accomplished by reading on a Luminex FlexMap 3-D system with a minimum of 50 bead reads per well. Median fluorescence intensity (MFI) values were reported for the conditions to permit the evaluation of coupling efficiency and preliminary evaluation of an assay range.

Preliminary evaluation of the assay demonstrates significantly higher circulating levels of anti-vimentin antibodies in 51 cases of renal allograft rejection relative to 20 cases of age-matched controls. From repeated testing of three distinct batches of assays provide assay range parameters of 0.18–15 μg/mL, median inter-assay recovery parameter was within 1% of completion, and interassay variation was at 7%.

The authors concluded that their direct capture assay for vimentin autoantibodies was successfully developed and analytically validated. Preliminary evaluation of this assay against patient materials was promising and justifies additional testing with larger cohorts in future studies. The study was published in the May 2014 issue of Journal of Immunological Methods.

Related Links:
Rush University Medical Center
Luminex Corporation
R&D Systems