# Sensitivity Enhancement of Multiplex Lateral Flow Immunoassays by NIR-II Fluorescence and Thermal Contrast

## TL;DR

This paper introduces a new lateral flow assay platform that uses advanced fluorescence and thermal signals to improve sensitivity and detect multiple cancer biomarkers at the point of care.

## Contribution

A multimodal LFA platform integrating NIR-II fluorescence and photothermal readouts for enhanced sensitivity and multiplex detection.

## Key findings

- The platform achieved comparable detection limits for CA15-3 (0.40 and 0.42 U/mL) using thermometric and NIR-II fluorescence modes.
- CEA was detected with a limit of 0.096 ng/mL, demonstrating high sensitivity.
- Multiplexed detection of CA15-3 and CEA on a single strip showed minimal cross-reactivity and strong correlation with standard assays.

## Abstract

Lateral flow assays (LFAs) are widely used for point-of-care
(POC)
diagnostics but often suffer from limited sensitivity and specificity
compared with laboratory methods. Here, we present a multimodal LFA
platform integrating colorimetric, photothermal, and second near-infrared
window (NIR-II) fluorescence readouts for enhanced sensitivity and
multiplexed detection. Gold nanorods were coupled with bright NIR-II
emissive polymer dots to generate plasmon-enhanced fluorescence and
efficient photothermal signals within a single probe. As proof-of-concept,
carbohydrate antigen 15-3 (CA15-3) and carcinoembryonic antigen (CEA)
were selected as target biomarkers for breast cancer screening and
broader cancer indication, respectively. Both thermometric and NIR-II
fluorescence modes achieved comparable limits of detection for CA15-3
(0.40 and 0.42 U/mL) with a dynamic range of 0–100 U/mL, while
CEA was quantified with a detection limit of 0.096 ng/mL. Multiplexed
analysis on a single strip allowed simultaneous detection of CA15-3
and CEA with minimal cross-reactivity, and NIR-II fluorescence from
test line 2 was intentionally designed to be invisible to the naked
eye to avoid interference with rapid CA15-3 screening. Validation
with clinical serum samples demonstrated a strong correlation with
standard electrochemiluminescence immunoassays. This portable, low-cost
platform demonstrates that NIR-II fluorescence and photothermal readouts
can be harmonized for sensitive, selective, and multiplexed POC cancer
biomarker detection. This universal and signal-amplifying concept
can be further adapted to other targets of interest, offering a promising
route toward next-generation LFAs.