Detection Of Rabbit IgG Research Articles

Copper deposition-induced efficient signal amplification for ultrasensitive lateral flow immunoassay

This work describes a novel signal amplification strategy via gold nanoparticle (AuNP)–induced copper deposition to develop a highly sensitive and accurate immunochromatographic platform. Copper deposition–assisted signal amplification (CDASA) was first evaluated on commercially HCG test strip. A lower limit of detection (LOD) over the well-known silver enhancement confirms the great potential of this new approach for the visual test strip. Under an optimal condition, an LOD of 1 pg mL−1 and a wide dynamic range from 1 pg mL−1 to 100 ng mL−1 were achieved by the CDASA with rabbit IgG as a model analyte. In terms of LOD, 2–3 orders of magnitude improvement is received by the CDASA as compared with the conventional test strip without any signal amplification. Potential of the proposed strategy for practical application is also validated by detection of Rabbit IgG in human serum, a mimic clinical sample. The CDASA renders much superiority such as a higher deposition efficiency, lower cost, and better reagent stability over the silver enhancement method. The results can be qualitative and quantitative analysis by the naked eyes and the combination of a flatbed scanner and a free imageJ respectively, eliminating any lab equipment.

Detection of rabbit IgG by using functional magnetic particles and an enzyme-conjugated antibody with a homemade magnetic microplate.

BackgroundThe enzyme-linked immunosorbent assay (ELISA) has been used for diagnosing medical and plant pathologies. In addition, it is used for quality-control evaluations in various industries. The ELISA is the simplest method for obtaining excellent results; however, it is time consuming because the immunoreagents interact only on the contact surfaces. Antibody-labeled magnetic particles can be dispersed in a solution to yield a pseudohomogeneous reaction with antigens which improved the efficiency of immunoreaction, and can be easily separated from the unreactive substances by applying a magnetic force. We used a homemade magnetic microplate, functional magnetic particles (MPs) and enzyme-labeled secondary antibody to perform the sandwich ELISA successfully.ResultsUsing antibody-labeled MPs enabled reducing the analysis time to one-third of that required in using a conventional ELISA. The secondary antibody conjugated with horseradish peroxidase (HRP) was affinity-bound to the analyte (IgG in this study). The calibration curve was established according to the measured absorbance of the 3, 3′, 5, 5′-tetramethybezidine–HRP reaction products versus the concentrations of standard IgG. The linear range of IgG detection was 114 ng/mL–3.5 ng/mL. The limit of detection (LOD) of IgG was 3.4 ng/mL. The recovery and coefficient of variation were 100% (±7%) and 116% (±4%) for the spiked concentrations of 56.8 ng/mL and 14.2 ng/mL, respectively.ConclusionPseudohomogeneous reactions can be performed using functional MPs and a magnetic microplate. Using antibody-labeled MPs, the analysis time can be reduced to one-third of that required in using a conventional ELISA. The substrate–enzyme reaction products can be easily transferred to another microplate, and their absorbance can be measured without interference by light scattering caused by magnetic microbeads. This method demonstrates great potential for detecting other biomarkers and in biochemical applications.Graphical A magnetic ELISA with convenient magnetic microplate.Electronic supplementary materialThe online version of this article (doi:10.1186/s13065-015-0088-1) contains supplementary material, which is available to authorized users.

Highly Stable Porous Silicon–Carbon Composites as Label-Free Optical Biosensors

A stable, label-free optical biosensor based on a porous silicon-carbon (pSi-C) composite is demonstrated. The material is prepared by electrochemical anodization of crystalline Si in an HF-containing electrolyte to generate a porous Si template, followed by infiltration of poly(furfuryl) alcohol (PFA) and subsequent carbonization to generate the pSi-C composite as an optically smooth thin film. The pSi-C sensor is significantly more stable toward aqueous buffer solutions (pH 7.4 or 12) compared to thermally oxidized (in air, 800 °C), hydrosilylated (with undecylenic acid), or hydrocarbonized (with acetylene, 700 °C) porous Si samples prepared and tested under similar conditions. Aqueous stability of the pSi-C sensor is comparable to related optical biosensors based on porous TiO(2) or porous Al(2)O(3). Label-free optical interferometric biosensing with the pSi-C composite is demonstrated by detection of rabbit IgG on a protein-A-modified chip and confirmed with control experiments using chicken IgG (which shows no affinity for protein A). The pSi-C sensor binds significantly more of the protein A capture probe than porous TiO(2) or porous Al(2)O(3), and the sensitivity of the protein-A-modified pSi-C sensor to rabbit IgG is found to be ~2× greater than label-free optical biosensors constructed from these other two materials.

Magnetic and electrokinetic manipulations on a microchip device for bead‐based immunosensing applications

The combination of electrophoretic and magnetic manipulations with electrochemical detection for a versatile microfluidic and bead-based biosensing application is demonstrated. Amperometric detection is performed in an off-channel setup by means of a voltammetric cell built at the microchannel outlet and using a gold working electrode. Superparamagnetic particles are introduced and handled inside the channel by means of an external permanent magnet in combination with the electrogenerated flow which allows reproducible loading. The specific detection of phenol as electroactive alkaline phosphatase product is used in this study as proof of concept for a sensitive protein quantification. Characterizations and optimization of different parameters have been carried out in order to achieve the best detection signal. The applicability of the device has been finally demonstrated by the detection of rabbit IgG as model protein after an immunoassay performed on magnetic particles as immobilization platform. A comparison between the electrochemical detection using the developed device and the optical standard detection revealed similar performances with, however, extremely lower amount of reagent used and shorter analysis time. The developed electrophoretic- and magnetic-based chip may open the way to several other biosensing applications with interest not only for other proteins but also for DNA analysis, cell counting, and environmental control.

Paper‐Based ELISA

Paper works: Paper-based indirect ELISA (see picture) has been demonstrated through the detection of rabbit IgG and the HIV-1 envelope antigen gp41. This technique combines the sensitivity and specificity of ELISA with the low cost and ease-of-use of paper-based platforms.

Pre-screening for antigen detectability in cells: a TEM-based solid phase digital immunogold detection method utilizing ultra low volumes of reagents.

Rapid and sensitive pre-screening for the presence of antigens in cell samples and confirmation of reactivity of antibodies, before proceeding with electron microscopy, is highly desirable. Most of the methods developed for this purpose are generally not very efficient and suitable for dealing with very small volumes of sample and reagents. In this work we present a simple, sensitive and rapid solid phase transmission electron microscope (TEM) based method for the detection of picogram (pg) levels of soluble antigens using as little as 10 micro L of reagents. Protein was adsorbed onto grids coated with polystyrene films to form the solid phase. The presence of antigen was detected using immunogold labelling. Gold particles adhering to the film were visualized and counted in a TEM providing a digital signal. This method was 100-fold more sensitive than dot blot in detection of rabbit IgG. We have demonstrated the utility of this technique by screening for Vitreoscilla haemoglobin (VHb) antigen in cell lysates and confirming the results directly with immunogold labelling transmission electron microscopy of cell sections.