# A multiplexed micro scale assay for real time analysis of pediatric immune cell function

> **NIH NIH U24** · UNIVERSITY OF WISCONSIN-MADISON · 2022 · $360,396

## Abstract

Abstract
Restricted amounts of biological samples that can be collected from pediatric patients
for the study of immune function are often insufficient for conventional assays.
Development of novel, multi-parameter sample sparing assays is necessary for
advancing the fundamental understanding of the immune system. In this proposal, we
outline three aims which detail the addition and integration of new biological readouts to
an existing microfluidic technology and the use of this expanded platform to investigate
the mechanisms for how allergies and asthma develop in children. The kit-on-a-lid-
assay (KOALA) is an easy to use self-contained microfluidic platform which is capable
of isolating neutrophils from whole blood, and assessing their chemotactic abilities. In
Aim 1 we propose to adapt and validate assays for measuring neutrophil ROS
production, NETosis, phagocytosis, cytokine production, and gene expression (RT-
qPCR) into KOALA. We also plan to modify the cell capture specificity of KOALA to
allow us to isolate T cells from whole blood and assess their cytokine production and
gene expression (RT-qPCR). The last component of Aim 1 is to test and compare a
new, less invasive blood draw method called Hemolink™ against traditional blood draw
methods in terms of immune cell phenotype. In Aim 2 we propose to use a single-cell
real-time non-invasive label-free microscopy method called optical metabolic imaging
(OMI) to identify single-cell heterogeneity in activation within T cell and neutrophil
populations. OMI is compatible with the other endpoints and can be run simultaneously.
In Aim 3 will apply our multiplexed KOALA assays to blood samples from a cohort of
children with allergic disease and matched healthy controls from the WISC cohort (an
ongoing study at UW that includes farm and non-farm children). We will compare our
neutrophil and T cell function and phenotype results from these children with the
extensive measurements being done as part of the WISC cohort study. In Aim 3 we will
also extend these analyses to a group of infants from the CREW study (a new study at
UW that aims to identify early life genetic and environmental risk factors for allergic
diseases and asthma) using the HemoLink™ device to draw blood with minimal
discomfort and profile type 2 T cell function (T cells differentiated to secrete IL-4, IL-5,
IL-13 and other type 2 cytokines) using KOALA. If successful in our aims, we will have
created a miniaturized, highly multiplexed, self-contained microfluidic platform that is
capable of measuring biological markers related to immune status and function. This
system will have then been validated in pediatric subjects and ready for transition into
clinical research.

## Key facts

- **NIH application ID:** 10380807
- **Project number:** 5U24AI152177-03
- **Recipient organization:** UNIVERSITY OF WISCONSIN-MADISON
- **Principal Investigator:** David J Beebe
- **Activity code:** U24 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $360,396
- **Award type:** 5
- **Project period:** 2020-04-01 → 2025-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10380807

## Citation

> US National Institutes of Health, RePORTER application 10380807, A multiplexed micro scale assay for real time analysis of pediatric immune cell function (5U24AI152177-03). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10380807. Licensed CC0.

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