A Magnetic Nanoparticle-Based Immunoassay Using a Microfluidic Device

Abstract

Source: Hernández-Ortiz, J. A., et al. Computer Numerical Control Micromilling of a Microfluidic Acrylic Device with a Staggered Restriction for Magnetic Nanoparticle-Based Immunoassays. J. Vis. Exp. (2022).

This video demonstrates a microfluidic system for a magnetic nanoparticle-based immunoassay. The detection system utilizes a microfluidic device incorporating a magnetic trap within its channels. Pre-formed immunocomplexes, comprised of antigen-coated magnetic nanoparticles bound to primary and peroxidase-conjugated secondary antibodies, are introduced into the channels. The immunocomplexes become captured in the porous magnetic trap upon applying an external magnetic field. Using a fluorogenic substrate emitting fluorescence upon catalysis by the peroxidases, the captured immunocomplexes are detected under a microscope.

Protocol

1. Device preparation

1. Fill the channels with distilled water using a syringe. Make sure there are no leaks or resistance to flow. Immerse the device in an ultrasonic bath for 10 min to remove any remaining acrylic, adhesive, or unwanted material inside the channels.
2. Empty the water inside the device channels. Use a syringe to introduce a blocking solution prepared with 5% (w/v) bovine serum albumin (BSA) diluted in 1x Tris-buffered saline (TBS) and previously filtered through a 0.2 µm polyethersulfone (PES) syringe filter.
3. Prepare a suspension of iron microparticles of 7.5 µm diameter in 5% BSA.
   

Results

Figure 1: Final device configuration. (A) Acrylic device with the hoses attached to the corresponding inputs and outputs. The scale shows the dimensions of the device in centimeters. (B) Protocol for the formation of .......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Carbonyl-iron microparticles	Sigma-Aldrich	44890	7 μm
Chloroform	Fermont	6201	Health Hazard: Moderate Flammability: None Reactivity: None Contact Hazard: Moderate
CMOS camera Moment	Teledyne Photometrics		Sensor Technology: CMOS Quantum Efficiency: 73% Pixel Size: 4.5 µm x 4.5 µm Supported Interfaces: USB 3.2 Gen 2
Dr Engrave Software	Roland DGA Corporation		Engraving software to design and create the engraving path on the surface
Extraction hood	Unknown	Unknown
Flexible Plastic Tubing	Tygon	AAD04103	ID = 0.020, OD = 0.060
Fluorescence microsope	ZEISS	Axio Vert.A1
High Precision Dispense Needle	Loctite	98612
MagJET Separation Rack	thermoscientific		12 x 1.5 mL
Polymethylmethacrylate - Sheet - PMMA, Acrylic	Goodfellow	ME303018/1	Thickness: 1.3 mm, Transparency: Clear/Transparent
PVCamTest software	Teledyne Photometrics	Version 3.10.107	Image acquisition software
Stereo microscope	Nikon	SMZ 7457
SuperMag Carboxyl Beads	Ocean NanoTech	KSC0100	100 nm
Syringe pump	kd Scientific	KDS200	Can hold up to two syringes
Utrasonic bath	Branson	2800
VPanel software	Windows OS	Version 1.0.3.0	Software for controlling the micromilling machine