研究目的
Developing a highly sensitive and selective method for detecting human cardiac troponin-I (cTnI) in complex serum samples using immunomagnetic separation technology-assisted surface plasmon resonance (SPR) biosensing.
研究成果
The developed method significantly enhances the sensitivity of SPR biosensing for cTnI detection through immunomagnetic separation and signal amplification, achieving a 320-fold lower detection limit compared to PDA-based methods. It shows high specificity, good accuracy in serum samples, and potential for clinical diagnosis. Future work could focus on improving instrumental sensitivity and expanding the method to other proteins.
研究不足
The detection limit (3.75 ng mL-1) may not be as low as some other reported methods due to instrumental limitations of the lab-built SPR biosensor. The reusability of the sensing platform decreases after multiple uses (78% activity after fourth use).
1：Experimental Design and Method Selection:
The study employs a combination of immunomagnetic separation and SPR biosensing. Fe3O4@PDA nanoparticles are used as magnetic immune probes for target capture and enrichment, and MWCNTs-PDA-AgNPs are used for signal amplification. The sensing platform is modified with PDA and AuNPs for antibody immobilization.
2：Sample Selection and Data Sources:
Human serum samples spiked with cTnI are used. Samples are provided by the Second Hospital of Jilin University and stored at -20°C.
3：List of Experimental Equipment and Materials:
Key materials include Fe3O4 nanoparticles, dopamine, multi-walled carbon nanotubes, antibodies (cAb, dAb, Ab2), HAuCl4, BSA, Tris buffer, PBS. Equipment includes TEM, SEM, XPS, VSM, UV-vis spectrometer, and a lab-built SPR biosensor.
4：Experimental Procedures and Operational Workflow:
Synthesis of Fe3O4@PDA-dAb immune probes, preparation of MWCNTs-PDA-AgNPs/Ab2 nanoconjugates, modification of the Au film sensing platform with PDA and AuNPs, immobilization of cAb, magnetic separation of cTnI from samples using immune probes, injection of concentrated samples onto the sensing platform, and secondary signal amplification with MWCNTs-PDA-AgNPs/Ab2. SPR spectra are recorded.
5：SPR spectra are recorded. Data Analysis Methods:
5. Data Analysis Methods: SPR response signals (resonant wavelength shifts) are measured. Detection limit is defined as the lowest concentration with a signal three standard deviations above the blank. Optimization of parameters (e.g., mass ratio of dAb to Fe3O4@PDA, concentration of immune probes) is done using UV-vis spectroscopy and SPR measurements.
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Fe3O4 nanoparticles
Used as magnetic cores for immune probes to enable magnetic separation and enrichment of target analytes.
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Multi-walled carbon nanotubes
Outer diameters of 30–50 nm, lengths of 0.5–2 μm
Chengdu Organic Chemicals Co., Ltd.
Used as supports for dispersing Ag nanoparticles in nanocomposites for signal amplification.
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SPR biosensor
Lab-built wavelength modulation SPR biosensor
Used for recording SPR spectra and measuring resonant wavelength shifts in biosensing experiments.
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Transmission electron microscope
Used for characterizing the core-shell structure of nanoparticles.
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Scanning electron microscope
Used for imaging the surface morphology of modified films and nanocomposites.
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X-ray photoelectron spectrometer
Used for analyzing the elemental composition of nanoparticles.
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Vibrating sample magnetometer
Used for evaluating the magnetic properties of nanoparticles.
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UV-vis spectrometer
Used for measuring absorption spectra to verify conjugation and optimize parameters.
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Millipore purification device
Millipore
Used for preparing deionized water for experiments.
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