Biomedical Microdevices

, Volume 13, Issue 3, pp 573–583 | Cite as

Microfluidic pillar array sandwich immunofluorescence assay for ocular diagnostics

  • James V. Green
  • Dawei Sun
  • Ali Hafezi-Moghadam
  • Kameran Lashkari
  • Shashi K. Murthy
Article

Abstract

Uveitis and primary intraocular lymphoma (PIOL) are diseases associated with the invasion of lymphocytes into various regions of the eye, accompanied by expression of inflammatory cytokines. While these diseases are very different in terms of survivability and treatment options they have similar symptoms that make accurate diagnosis challenging. Furthermore, the diagnostic yield with state-of-the-art techniques for cell and cytokine analysis of vitreous and aqueous humor samples is under 20% due to inadequate sensitivity. This paper describes a simple sandwich immunofluorescence assay (sIFA) microfluidic device that is capable of identifying important analytes in ocular biopsies as a potential alternative to current diagnostic approaches. Detection is accomplished by capture of the target molecules on antibody-coated, vertical, oval shaped pillars in a microfluidic device followed by a biotinylated detection antibody and finally florescent avidin for target molecule quantification. Cytokine concentration measurements were carried out on aqueous humor samples from rats with endotoxin-induced uveitis as well as human cataract patients. Results correlated well with conventional protein quantification techniques and additionally, measurements from the human samples surpassed detection limits of current state-of-the-art immunoassay techniques. The single-digit femtomolar range of detection of this sIFA system provides lower limits of detection when compared to traditional techniques and allows for the mapping of the cytokine content of vitreous biopsies with detection limits that have yet to be realized using cost effective microfluidics. Furthermore, the relative simplicity of the device design, fabrication and ability to automate makes it easily translatable from the laboratory to a clinical setting.

Keywords

Microfluidic Immunoassay Uveitis Primary intraocular lymphoma 

Supplementary material

10544_2011_9528_MOESM1_ESM.doc (128 kb)
Fig. S1Rat cytokine calibration curves for (a) TNFα, (b) IL-6 and (c) IL-10 (DOC 128 kb)
10544_2011_9528_MOESM2_ESM.doc (334 kb)
Fig. S2Natural log plots of the rat cytokine calibration curves (a) TNFα, (b) IL-6 and (c) IL-10. (DOC 333 kb)
10544_2011_9528_MOESM3_ESM.doc (121 kb)
Fig. S3Human cytokine calibration curves (a) TNFα, (b) IL-5 and (c) IL-10 (DOC 121 kb)
10544_2011_9528_MOESM4_ESM.doc (359 kb)
Fig. S4Natural log plots of the human cytokine calibration curves (a) TNFα, (b) IL-5 and (c) IL-10 (DOC 359 kb)

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Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • James V. Green
    • 1
  • Dawei Sun
    • 2
    • 4
  • Ali Hafezi-Moghadam
    • 2
    • 4
  • Kameran Lashkari
    • 3
    • 4
  • Shashi K. Murthy
    • 1
  1. 1.Department of Chemical EngineeringNortheastern UniversityBostonUSA
  2. 2.Department of OphthalmologyMassachusetts Eye and Ear InfirmaryBostonUSA
  3. 3.Schepens Eye Research InstituteBostonUSA
  4. 4.Harvard Medical SchoolBostonUSA

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