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Time：2020-08-17Click：
Current and future applications of colloidal gold for rapid detection
James Carney, Helen Braven, Joanna Seal, and Emma Whitworth
Rainylily comrades translation
Over the past five years, the use of gold particles and chromatography has increasingly established its role in bedside detection. The advent of simple chromatography for different analytes simplifies and speeds up the detection system, which also has a great impact on many enterprises. For example, a drop of blood, urine or saliva from a patient can be used during a clinical interview to make an accurate diagnosis so that treatment can begin immediately while the patient is still present. Similarly, food production companies can use quality control testing at different stages of food processing without the need for dedicated laboratory staff or disruption of the production process. In addition, tomographic tests can be used in veterinary practice, food storage and environmental monitoring without the need for specialized equipment and training and interpretation of the results. Coupled with the rapid reaction, chromatography has become an ideal means of self-detection.
Improvements in the materials used to produce tomographic tests, as well as in the design of experiments, have increased the demand for more tomographic tests that can produce quantitative results. As the multi-billion dollar global in vitro diagnostic test market grows, these same drivers can also lead to the diversification of rapid tests and the convenience of laboratory analysis. This article discusses the causes of this diversification and Outlines some of the properties of colloidal gold for a variety of diagnostic applications.
The use of colloidal gold in chromatography detection
Two kinds of particles are often used for chromatography detection: stained latex and colloidal gold. Many recently developed chromatography tests, including Phadia AB's (Uppsala, Sweden) Rapid trial of Immunocap Rapid and Merck KgGA's (Darmstadt, Germany) Singlepath and Duopath trials for detecting food-borne pathogens, use nanogold as markers.
The size of gold nanoparticles for chromatography detection is smaller than that of latex particles, and the pore size of nitrocellulose films for detection is 8-10 microns, which can accommodate a large number of nanoparticles at a time. The sensitivity of this form of detection depends on the degree of mixing of the particle and analyte as they move along the membrane towards the adsorption line (see Figure 1). Another important factor is that small particles are more closely packed on the adsorption line and thus more easily identifiable.
FIG. 1 Schematic diagram of rapid chromatography detection
IDV manufacturers must purchase colloidal gold from reputable suppliers to ensure that strict quality control standards are met to achieve uniform particle size and size, which is also a major factor in maintaining free agglutination and stable storage of diagnostic products. Colloidal gold is a homogeneous solution of particles with uniform surface area and charge, satisfying the two main criteria for protein adhesion.
The size of colloidal gold particles used for chromatographic detection ranges from 5 to 200nm. The range of particle size makes it applicable to different fields. Smaller particles can be used in the life sciences.
In order to apply the strongly identifiable characteristics of large particles by chromatography, particle homogeneity is critical. High quality colloidal gold particles must be monodispersed, spherical, and the number of heterogeneous particles should be less than 5%. Other colloidal gold specifications will result in poor results and a waste of time and resources. High quality colloidal gold can yield more results per liter.
Colloidal gold with a recognisable range of 20-120nm emits green light and thus appears bright red. Even if the particle is smaller than the wavelength of the transmitted light it can still emit light due to surface plasmon resonance. The wavelength of visible light emitted varies with the size of the particles.
Unlike the covalent binding used in latex, proteins such as antibodies are passively bound to colloidal gold. This combination is a relatively simple process without the use of other reagents. There are three commonly used modes of gold particle binding.
Standard colloidal gold particles are coated with a layer of negatively charged ions that bind to the surface of all the proteins. When gold particles are made with citric acid, they bind to amino acids such as lysine.
Amino acids with high hydrophobicity, such as tyrosine and tryptophan, bind to the surface of particles such as gold by hydrophobicity. Hydrophobic binding plays an important role in proteins with high hydrophobic amino acid content, such as immunoglobulin, which is why the binding particles are less reactive after prolonged exposure to the detergent containing buffer.
A gold-sulfur bond is a strong bond formed when gold and sulfur share an electron pair. The attachment of the gold particle to the cysteine residues in the protein may be the most important part of the antibody or antigen that attaches to the particle. Therefore, the use of sulfur-containing buffers or preservatives such as thiomersal should be avoided.
Controllability through strict quality control procedures to produce a consistent size of the required round colloidal gold particles. Years of development experience have eliminated variation in mass production and the difficulty of mass supply (Figure 2). Colloidal gold is simply connected, involving only proteins, diluents and particles without the need for reagents. Based on the specified raw materials, IDV producers can calculate the exact number of antibodies required for each colloidal gold to make the best detection result, thus saving cost and reducing the waste of raw materials.
FIG. 2 High quality round nanometer gold has low variation coefficient, consistent morphology, and optimal antibody adsorption and sensitivity.
Improvements in the quality of raw materials used for chromatography detection have improved the performance of the detection.
Improvements in particle processing have led to the continuous production of large size (250nm) round colloidal gold particles, which can be used as a variety of markers on the market. The 40nm size particles are considered the most suitable for chromatography detection. This size is both large enough to be easily discernible and small enough not to cause steric hindration to the binding of the protein to the colloidal gold surface to optimize the performance of the labeled raw material. With the increase of particle size, its identifiability also increases. Initial studies have shown that the application of colloidal gold at 60nm can increase the identification of the terminal signals in some tests, thus potentially improving the sensitivity of the test.
An important feature of this cellulose nitrate film is that its pore size controls the effective surface area of protein binding and the capillary flow rate of the sample through the test strip. Poor quality colloidal gold tends to aggregate and cannot pass freely through cellulose membrane, thus colloidal
The comparison of size distribution data in figure 3 shows that the average diameter of the labeled colloidal gold at 40 nm is 143nm
The gold particles must be uniform in size. IDV manufacturers must also guarantee colloidal gold size specifications. The size distribution comparison shows that the production of coarse colloidal gold labeled as 40nm May contain particles of different sizes with an average diameter of up to 143nm, resulting in incorrect results (Figure 3). The new nitrocellulose film is made to be wettable and does not affect protein absorption and test function. The post-processing of the product with surfactants and some blockers can improve the fluidity of the particles and eliminate the non-specific interaction between the test components.
Sample pad Sample pad can be impregnated with chemicals to reduce the difference in sample composition and improve the detection sensitivity. Detergents, thickeners, blockers and salts can usually be added to the sample pad after drying. This process avoids the use of complex development/tracking buffers and achieves true one-step detection.
Other membranes Today's blood-separation membranes are efficient at separating unhemolytic red blood cells and plasma from veins or capillaries. Both dc and flow-measuring films have been marketed for the separation of 10-110ml whole blood samples, which allows direct detection of serum or plasma without sample treatment and centrifugation.
Nucleotide chromatography detection has been developed to detect nucleotides in a variety of forms. The measurement of
FIG. 4 The inferior colloidal gold of 40nm is non-circular in shape.
Non - uniform, easy to aggregate, high coefficient of variation between particles
Products of amplification techniques such as polymerase chain reaction (PCR) and helicobacter pyloride-dependent amplification can be detected by this method. Helicobacter pyloride-dependent amplification is an isothermal amplification method similar to PCR, in which DNA strand separation is accomplished by enzyme action rather than heating. These techniques can be tested without expensive equipment.
Figure 5. (a) As a primer for PCR amplification, double-stranded PCR or tHDA products were detected; (b) Detection of unlabeled single-stranded nucleic acid targets using sequence-specific oligonucleotide probes with fixed adsorptive antibodies, antibody-colloidal gold binders, and hapten labeled; (c) Antibody-free detection using fixed oligonucleotide adsorption probes and oligonucleotide binding colloidal gold.
Nonsequence-specific detection This is a method of detecting the presence or absence of nucleotide analytes in the form of chromatography and using antibodies or hapten markers for nucleotides. Using this method, the two haptens (DNP and biotin) capture and detect the analyte by binding to an anti-DNP antibody (fixed on the chromatography band) and an anti-biotin antibody (bound to a gold nanoparticle). Haptens are stable at high temperatures
It is advantageous and does not affect the activity of nucleotide modifying enzymes. Haptens can also be commercially produced by oligonucleotide synthesis. This method is suitable for the detection of double-chain amplification products using biotin and DNP-labeled reagents (Figure 5a).
Sequence-specific detection is required for many molecular diagnostic applications. This assay allows the sequence of nucleotide analytes to distinguish them from other analytes and non-specific amplification products. Sequence specificity was achieved by annealing the oligonucleotide probe with the target sequence. There are several ways to ensure that the signal generation is dependent on either fixing the target with a probe or by prop-target annealing with labeled gold nanoparticles (Figure 5B and 5C).
Figure 6. Oligonucleotide sequences were detected by fixed oligonucleotide adsorption probes and oligonucleotide - colloidal gold complexes. The fixed probes that detect different sequences form test strips on the nitrocellulose membrane, nucleic acids containing each sequence are adsorbed at various locations, and then detected with another probe that binds to colloidal gold to produce a set of spectral lines.
Antibody-based detection systems for antibody-based nucleotide layers can reduce costs and increase detection repeatability, sensitivity, and specificity. The antibody-based nucleotide chromatography detection system is based on target capture by immobilized oligonucleotide probes and detection by probes directly bound to nanometer gold (as shown in Figure 5C). Sequence-specific oligonucleotide probes can be immobilized on chromatography bands using a method similar to speckle detection. The sensitivity and specificity of the capture probe is comparable to that of the method using antibodies, and there are significant advantages in multiple analyses (such as using multiple adsorption lines on each strip) (Figure 6). Various techniques have been used to develop ways of binding nucleotides to nanomaterials to obtain stable and strong bonds.
Other applications of colloidal gold. Colloidal gold is also effective in rapid tests that require instrumentation to discern the final result. These tests take advantage of the light scattering properties of colloidal gold. Although colloidal gold at 20-120nm has a smaller wavelength than visible light, it is still effective in light scattering due to its surface plasmon resonance properties. Surface plasmon resonance (SPR) is the result of an interaction between a specific wavelength of projected light and conducting electrons present in nanomaterials. The intensity of light scattering depends on the wavelength of the projected light and the size of the particles. This surface plasmon resonance property can be developed as a method for detecting tagged particles.
Colloidal gold can also be used as a fluorescent labeling substitute in microdisplays due to its surface plasmon resonance properties. The sensitivity of detecting bacterial RNA with gold nanoparticles by resonance light scattering is 50 times higher than that by fluorescence labeling assay. Another advantage of using gold particles is that they do not cause photobleaching.
The intensity and maximum wavelength of light scattering are positively correlated with the size of gold nanoparticles. By mixing colloidal gold particles of different diameters and selecting different particles for use in different analytes, it will be possible to develop multiple detection methods, with the addition of distinguishing plasmon resonances for particles of different sizes. This expands the range of tests that can be applied to chromatography or other forms of detection.
The use of gold nanoparticles led Pointcare Technologies (Marlborough, MA) to develop a CD4 test using the principle of simple current cell counting. The developed test eliminates a variety of fluorescent markers and intricate detection measures. This CD4 lymphocyte count USES colloidal gold anti-CD4 antibody binding agents that specifically bind to the surface of these lymphocytes, which emit light in a specific direction that distinguishes them from other types of cells, such as monocytes that express CD4. This simple, small, portable cell counting device can also count white blood cells and lymphocytes. It can also be used to monitor the effectiveness of HIV and AIDS patients.
Application of colloidal gold in light scattering diagnosis
People have been very interested in the development of other light scattering technologies involving colloidal gold and nano silver. At present, the surface enhanced Raman spectroscopy technology (SERS) has been applied in a variety of fields, and the application of nano gold in SERS detection has also been increasingly extensive, especially in the field of biological analysis.
Raman spectroscopy (SERS) method Raman scattering refers to the scattering of the projected light at different wavelengths. Because each kind of substance has its unique Raman spectrum, makes SERS become an excellent identification tool.
However, Raman signals also have their own weaknesses: about 1 in 107 incident photons has a wavelength bias when scattering. These signals can be enhanced in two ways. One method is resonant Raman scattering, in which the laser is tuned to detect the absorption wavelength of the material; Another method is SERS, which requires the detected material to be close to the surface of metal (gold or silver nanoparticles), and the plasma resonance characteristic can enhance the signal amplification to 105-106 times. After fusion of the two methods, it is called surface-enhanced Resonance Raman Scattering, which is a very sensitive technology, so it can amplify the signal 1014 times, and can see single molecule detection.
The optical properties of using nano metals as SERS matrix colloidal gold and nano silver made them good surfaces and substrates for SERS detection. Nanometer metals are suitable for liquid phase detection and have been widely used in many biological detection fields. Particle size, shape, gap and other factors are critical to signal enhancement, so high quality particles must be used. The development of a stable detection system relies on high-quality nanoparticles to produce stable products that can interact with biomolecules. The replacement of spherical particles (such as triangle or rod-shaped) was closely related to the application of SERS in the future and was beneficial to its application.
SERS tags Although SERS can be used for non-sers