Quantification of Immune Mediator Levels in Human Mucosal Lining Fluid

Abstract

Source: Wolsk, H. M. et al., Noninvasive Sampling of Mucosal Lining Fluid for the Quantification of In Vivo Upper Airway Immune-mediator Levels. J. Vis. Exp. (2017)

This video demonstrates the quantification of immune mediator levels in neonatal human nasal mucosal lining fluid extract. An electrochemiluminescence-based multiplex assay system measures these mediators by evaluating the intensity of emitted light when they are excited.

Protocol

All procedures involving sample collection have been performed in accordance with the institute's IRB guidelines. 1. Experimental Setup Use sheets of filter paper (fibrous hydroxylated-polyester sheets, see the Table of Materials). Cut out strips 3 x 15 mm in size for a 1-month-old child and in an L-shape for older children and adults (5 x 20 x 10 mm (short arm of the L)). Insert one piece…

Materials

Fibrous hydroxylatedpolyester sheets	Accuwik Ultra	SPR0730	Filter paper
Milliplex Assay Buffer	Millipore	L-AB	Buffer
low-protein binding storage plates	Thermo Scientific	CLS8161	Plates
Protease Inhibitor	Roche	11873580001	Complete EDTA-free Protease Inhibitor Cocktail
Reader of multi-spot plates	Mesoscale	NA	Sector imager 6000
Assays	Mesoscale		Human 10-plex TH1/TH2 cytokine assay and 9-plex chemokine assay, and singleplex IL-17A, TGF-β1and TSLP. A description can be found online on www.mesoscale.com