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.
1. Device preparation
2. Microparticle trap formation
3. Immunoassay
4. Experimental mounting
5. Immunodetection
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 the microparticle trap. Microparticles flow through the channel by gravity when the device is placed in a vertical position. Microparticles are concentrated at the 5 µm restriction. Excess microparticles are easily removed by rotating the chip through the side channel. The chip is kept vertical to preserve the trap before immunoassay. (C) Microfluidic device mounted on a glass slide containing the magnet, on the stage of the inverted fluorescence microscope. The dispensing needle through which the reagents are added is observed, as well as the outlet hoses that connect to a syringe pump.
Figure 2: Nanoparticle separation. Using a commercial magnetic separator, 100 nm nanoparticles can be easily concentrated to perform the washing steps during the immunoassay. The pellet formed after 15 min is observed in the red circle.
The authors have nothing to disclose.
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 |
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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 |