A Multiplex Bead-Based Assay for Analyzing Cytokines in Human Tear Samples

Abstract

Source: Balne, P. K., et al. Bead Based Multiplex Assay for Analysis of Tear Cytokine Profiles. J. Vis. Exp. (2017)

This video illustrates a bead-based multiplex assay for analyzing cytokines in human tears. The method allows for the simultaneous analysis of multiple cytokines from a single sample, providing enhanced sensitivity for studying immune responses in various pathophysiological conditions.

Protocol

All procedures involving sample collection have been performed in accordance with the institute's IRB guidelines.

1. Tear Cytokine Analysis

1. Collection of tears:
   1. Ask the subject to sit comfortably on an examination chair and place his/her head against the headrest.
   2. Ask the subject to look up, carefully open the Schirmer strips (made of Whatman filter paper no. 41) place the rounded end of the strip on the inferior fornix of the eye, and instruct the subject to close the eyes for 5 min. While collecting the tears take great care to minimize ocular surface contact.

Results

Figure 1: Schematic flow diagram of tear cytokine profiling.

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Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Milliplex MAP human cytokine / chemokine magnetic bead panel -1 kit	Merck, USA	HCYTOMAG-60K-41
Flexmap 3D luminex instrument	Luminex Corp, Austin, TX, USA
xPonent software	Luminex Corp, Austin, TX, USA
RBXGenerator software	BIO-RAD, France
Bio-Plex Manager 6.1	BIO-RAD, France
Plate shaker	Corning, USA
TECAN Microplate Washer	Tecan, Switzerland	HydroSpeed
Pipettes	Mettler Toledo, CA, USA
1.5 ml microcentrifuge tube	Axygen, USA	MCT-150-C
Schirmer tear flow test strip	Eye Care and Cure, USA	101657
Flexmap 3D Calibration Kit	Luminex Corp, Austin, TX, USA	40-028
Flexmap 3D Verification Kit	Luminex Corp, Austin, TX, USA	40-029
Ocular examination chair