Colloidal Gold-based Immunochromatographic Strip Research Articles

The Strategic Alliance between Clinical and Molecular Science in the War against SARS-CoV-2, with the Rapid-Diagnostics Test as an Indispensable Weapon for Front Line Doctors.

Our work concerns the actual problem of spread of SARS- CoV-2 outbreak which requires fast and correct as possible answer. In current scenario, the need of rapid answer put away the imperative of proper methodology. We focus on the serogical immunoassay for diagnosis of Covid-19 as an important weapon not only for diagnostic purpose, but also for epidemiologic one. The right equilibrium between high speed, low cost and accuracy is obtained with easy-to-use decentralized point-of-care test as the colloidal gold-based immunochromatographic strip assay which detects IgM and IgG antibodies directed against SARS-CoV-2. As our aim is to evaluate the efficacy of Covid-19 rapid tests and of serological assays in real-life settings, we designed a research protocol aimed to establish how to use correctly these diagnostics, taking into account the different possible clinical and epidemiological scenarios.

Colloidal gold-based immunochromatographic strip assay for the rapid detection of bacitracin zinc in milk

In this study, a murine monoclonal antibody (mAb) of 6D2-G10 against bacitracin zinc (BAC) was produced and applied to an immunochromatographic strip (ICS) for the initial detection of BAC in milk. The ICS with a cut-off value of 25 ng/mL could be perceived by the naked eye within 10 min. With the assist of the strip reader, the limit of detection (LOD) was measured as 0.82 ng/mL, the half-maximal inhibitory concentration (IC50) was recorded as 3.16 ng/mL, and the linear detection range was from 0.97 to 10.30 ng/mL. The recoveries ranged from 87.7% to 96.0% with the highest coefficient of variation (CV) of 9.1% in the intra-assay and from 84.3% to 90.2% with the highest CV of 10.7% in the inter-assay. In short, the established ICS provided a serviceable analytical tool for qualitatively and quantitatively monitoring BAC in milk.

Serological immunochromatographic approach in diagnosis with SARS-CoV-2 infected COVID-19 patients

An outbreak of new coronavirus SARS-CoV-2 was occurred in Wuhan, China and rapidly spread to other cities and nations. The standard diagnostic approach that widely adopted in the clinic is nucleic acid detection by real-time RT-PCR. However, the false-negative rate of the technique is unneglectable and serological methods are urgently warranted. Here, we presented the colloidal gold-based immunochromatographic (ICG) strip targeting viral IgM or IgG antibody and compared it with real-time RT-PCR. The sensitivity of ICG assay with IgM and IgG combinatorial detection in nucleic acid confirmed cases were 11.1%, 92.9% and 96.8% at the early stage (1–7 days after onset), intermediate stage (8–14 days after onset), and late stage (more than 15 days), respectively. The ICG detection capacity in nucleic acid-negative suspected cases was 43.6%. In addition, the concordance of whole blood samples and plasma showed Cohen's kappa value of 0.93, which represented the almost perfect agreement between two types of samples. In conclusion, serological ICG strip assay in detecting SARS-CoV-2 infection is both sensitive and consistent, which is considered as an excellent supplementary approach in clinical application.

Development of a colloidal gold-based immunochromatographic strip test using two monoclonal antibodies to detect H7N9 avian influenza virus.

H7N9 low pathogenic avian influenza viruses (AIVs) emerged in China in 2013 and mutated into highly pathogenic strains in 2017, causing disease in infected birds and humans. Thus, the development of rapid, specific, and sensitive detection methods is urgently required. Herein, two specific monoclonal antibodies against H7N9 AIV were produced to develop a colloidal gold-based immunochromatographic test strip to detect H7N9 AIV. High specificity, repeatability, and sensitivity were achieved, with a detection limit of two hemagglutinin units or 102.55 50% tissue culture infective dose. This assay may represent a powerful tool to rapidly detect H7N9 influenza viruses in the future.

One-step rapid detection of fumonisin B1, dexyonivalenol and zearalenone in grains

Mycotoxins are a class of secondary metabolites produced by filamentous fungi that are harmful to humans and animals. Here, a multiplex immunochromatographic assay was reported, which is based on 25 nm colloidal gold for simultaneous qualitative detection of fumonisin B1 (FB1), deoxynivalenol (DON) and zearalenone (ZEN) in wheat and corn samples. After conditions optimization, the immunochromatographic strip has high sensitivity. The visual detection limits of the immunochromatographic test strips for FB1, DON, and ZEN reached 60, 12.5 and 6 ng/mL, respectively. In addition, the immunochromatographic strip has high specificity, no reaction to each other, or no cross-reactivity with ochratoxin A (OTA) and aflatoxin B1 (AFB1). Furthermore, eighteen samples (including corn and wheat) were tested using the test strips and verified by liquid chromatography-mass spectrometry (LC-MS/MS). The results of strips were found to be consistent with those of liquid chromatography-mass spectrometry (LC-MS/MS). In short, this work demonstrates that the spherical colloidal gold-based immunochromatographic test strip we developed can be used to quickly and simultaneously monitor the existence of multiple mycotoxins in agricultural food samples.

Rapid quantitative determination of fentanyl in human urine and serum using a gold-based immunochromatographic strip sensor.

Fentanyl is a typical opioid that is used in surgical anesthesia. However, when abused, fentanyl can lead to addiction and even death. To better control the use of fentanyl, it is necessary to develop rapid and sensitive detection methods. In this study, an ultrasensitive monoclonal antibody (mAb) was prepared and used to develop an indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and a colloidal gold-based immunochromatographic strip (CG-ICS) for the analysis of fentanyl in urine and serum. Under optimum conditions, the anti-fentanyl mAb belonging to the subtype of IgG2b showed a half-maximal inhibitory concentration (IC50) of 0.11 ng mL-1 and a linear range of detection of 0.020-0.50 ng mL-1. Fenanyl-spiked original urine and serum diluted eight times were used for the analysis of fentanyl by ic-ELISA and CG-ICS. IC50 from the standard curves was 0.46 ng mL-1 for urine and 2.6 ng mL-1 for serum in ic-ELISA and 1.6 ng mL-1 for urine and 6.27 ng mL-1 for serum in CG-ICS. The recovery test revealed that the ic-ELISA and CG-ICS, with a recovery rate of 87.0-108.4% and a coefficient of variation of 3.3-10.9%, were the same reliable tools as the liquid chromatography tandem mass spectrometry for fentanyl analysis in real samples.

Development of a colloidal gold-based immunochromatographic strip for rapid detection of Rice stripe virus.

Rice stripe virus (RSV) causes dramatic losses in rice production worldwide. In this study, two monoclonal antibodies (MAbs) 16E6 and 11C1 against RSV and a colloidal gold-based immunochromatographic strip were developed for specific, sensitive, and rapid detection of RSV in rice plant and planthopper samples. The MAb 16E6 was conjugated with colloidal gold and the MAb 11C1 was coated on the test line of the nitrocellulose membrane of the test strip. The specificity of the test strip was confirmed by a positive reaction to RSV-infected rice plants and small brown planthopper (SBPH), and negative reactions to five other rice viruses, healthy rice plants, four other vectors of five rice viruses, and non-viruliferous SBPH. Sensitivity analyses showed that the test strip could detect the virus in RSV-infected rice plant tissue crude extracts diluted to 1:20 480 (w/v, g/mL), and in individual viruliferous SBPH homogenate diluted to 1:2560 (individual SPBH/μL). The validity of the developed strip was further confirmed by tests using field-collected rice and SBPH samples. This newly developed test strip is a low-cost, fast, and easy-to-use tool for on-site detection of RSV infection during field epidemiological studies and paddy field surveys, and thus can benefit decision-making for RSV management in the field.

Indirect competitive ELISA and colloidal gold-based immunochromatographic strip for amantadine detection in animal-derived foods

In this research, an indirect competitive enzyme-linked immunosorbent assay (ELISA) and colloidal gold-based immunochromatographic (GICG) strip were developed for the detection of the antiviral drug amantadine (AM) in animal-derived foods.

Development of colloidal gold‐based immunochromatographic strip test using two monoclonal antibodies for detection of Vibrio parahaemolyticus

AbstractA rapid and sensitive colloidal gold‐based immunochromatographic test strip (ICTS) for the detection of Vibrio parahaemolyticus (VP) was developed. It was based on sandwich principle with two monoclonal antibodies (Mabs), namely H7 and C9. The Mab H7 gold nanoparticle (AuNPs‐Mab H7) conjugate was transferred onto a glass fiber pad as conjugate pad. Mab C9 containing 1% (M/V) bovine serum albumin and the goat anti‐mouse immunoglobulinG (IgG) antibody were sprayed onto a nitrocellulose membrane in strips at T and C line positions, respectively. The ICTS was assessed for the ability to detect VP, 50 μl aliquot of test sample was applied to the sample pad. The results were observable within 10 min after adding the sample. The sensitivity of the test was determined to be 104 CFU/ml. Detection of VP after 25‐week storage could be accomplished with similar sensitivities. The positive rate was 100% in seafood with the ICTS. The VP ICTS provided several advantages over other detection methods, including the short time, low cost, convenient to use, and not requiring sophisticated equipment. Furthermore, the results could be imaged and transformed into a two‐dimensional bar code by using a smart cell phone for more vividly, clearly and permanently being observed.Practical applicationsDuring the past two decades, Vibrio parahaemolyticus (VP) has been one of the most common causes of the bacterial foodborne outbreaks, which caused abdominal cramps, vomiting, nausea, headaches, fever, and chills. Therefore, the detection of VP is crucial to public food safety. This study obtained two different monoclonal antibodies (Mabs; Mab H7 and C9) of VP with high specificity and no cross‐reactions were found with other foodborne pathogens. The AuNPs‐based immunochromatographic test strip (ICTS) was developed with the Mabs (Mab H7 and C9) and the ICTS was evaluated with high sensitivity, high specificity, low cost, and rapid speed for VP detection. The accuracy rate of immunochromatographic strip for detection VP was 100% in seafood samples.

Colloidal gold-based immunochromatographic strip assay for the rapid detection of three natural estrogens in milk

In this study, we developed highly sensitive and specific monoclonal antibodies (mAbs) against estrone (E1), 17β-estradiol (17β-E2), and estriol (E3). The half-maximal inhibitory concentration values of anti-E1, anti-17β-E2, and anti-E3 mAbs were 0.46, 0.36, and 0.39 ng/mL, respectively, based on competitive enzyme-linked immunosorbent assay (ic-ELISA) results. A rapid colloidal gold-based immunoassay strip assay was developed for the determination of E1, 17β-E2, and E3 residues in milk samples. The assay had a visual cut-off value of 5 ng/mL, and required 10 min to assess with the naked eye. The results obtained from the immunochromatographic strip assay were consistent with those obtained from ic-ELISA and gas chromatography–mass spectrometry. The immunochromatographic strip assay is useful and rapid for the detection of E1, 17β-E2, and E3 in milk.

Development of an immunochromatographic lateral flow strip for the simultaneous detection of aminoglycoside residues in milk.

A colloidal gold-based immunochromatographic strip with a competitive format has been developed for the rapid, simultaneous, semi-quantitative and quantitative detection of several aminoglycoside residues in milk, including gentamicin sulfate (GM), neomycin sulfate (NEO) and kanamycin sulfate (KN). Three monoclonal antibodies against the three corresponding aminoglycosides were conjugated to colloidal gold particles and applied to the conjugate pads of the strip. The competitors [GM-bovine serum albumin (GM-BSA), NEO-BSA and KN-BSA conjugates] of GM, NEO and KN were immobilized onto a nitrocellulose (NC) membrane at three detection zones, T1, T2, and T3, respectively. The minimal cut-off values of the strip were 10 ng mL−1 for GM, and 100 ng mL−1 for NEO and KN, which are lower than the maximum residue levels (MRLs) established for aminoglycosides. The IC50 values of the strip were 0.737 ng mL−1, 8.971 ng mL−1 and 11.110 ng mL−1 for GM, NEO and KN respectively. In conclusion, the immunochromatographic lateral flow strip could be used for rapid, simultaneous, semi-quantitative and quantitative detection of GM, NEO and KN residues in milk.

Lanthanide-Labeled Immunochromatographic Strip Assay for the On-Site Identification of Ancient Silk.

The on-site identification of ancient silks has long been a key challenge in archeology. Therefore, a rapid, cost-effective, sensitive analytical approach is highly desirable. In this paper, a lanthanide-labeled immunochromatographic strip which is suitable for the on-site identification of ancient silks is described. Compared with the conventional colloidal gold-based immunochromatographic strip, this strip shows much higher analytical sensitivity and better quantitative discrimination. The limit of detection (LOD) of the strip for silk fibroin (SF) was calculated as 8.09 ng/mL, approximately 185 times lower than that of the colloidal gold-based immunochromatographic strip. No cross-reactions with other possible interfering antigens were observed. Moreover, the strip also shows good reproducibility, with a mean recovery of 94.15-102.55% and coefficient of variation of 5.22-17.57% in the repeated tests. Based on the advantages of portability and cost-effectiveness, as well as sensitivity, specificity, and reproducibility, the lanthanide-labeled immunochromatographic strip is a promising tool for on-site detection of ancient relics in archeological fieldwork.

Urinary kidney injury molecule‑1 as an early diagnostic biomarker of obstructive acute kidney injury and development of a rapid detection method.

The aim of the present study was to investigate whether urinary kidney injury molecule-1 (KIM-1) presents a suitable early diagnostic biomarker of obstructive nephropathy-induced acute kidney injury (AKI), and to develop a rapid detection method for urinary KIM-1. Obstructive AKI was induced in an experimental rat model by a unilateral ureteral obstruction (UUO) operation. Macro- and micromorphological kidney alterations were determined by visual observation and hematoxylin and eosin (HE) staining, respectively. Kidney functions were evaluated by detecting urea nitrogen and creatinine levels in rat urine and blood. Urinary KIM-1 levels were measured using an enzyme-linked immunosorbent assay, and the protein expression levels of KIM-1, α-smooth muscle actin (α-SMA) and vimentin in kidney tissues were detected using immunohistochemical assays. In order to measure KIM-1 levels, colloidal gold immunochromatographic strips were developed based on the colloidal gold immunochromatographic assay. The results indicated that KIM-1 levels were significantly higher in the UUO group when compared with the Sham group. KIM-1 levels in the urine and kidney tissues exhibited a time-dependent increase, together with increasing obstructive AKI in the UUO group. In addition, KIM-1 levels were demonstrated to be a more sensitive biomarker of early obstructive AKI, when compared with α-SMA and vimentin. A colloidal gold-based immunochromatographic strip was developed, whereby the detection of urinary KIM-1 could be completed within 5–10 min. In conclusion, results of the present study demonstrated that urinary KIM-1 may be a valuable biomarker for the early diagnosis of obstructive AKI, and the use of a colloidal gold immunochromatographic strip may be a promising method for the rapid detection of urinary KIM-1.

An Immunochromatographic Test Strip to Detect Ochratoxin A and Zearalenone Simultaneously.

Lateral flow immunochromatographic assays are simple, useful tools for the detection of many different targets. In this format, a liquid sample containing target of interest moves along a strip of polymeric material adhered with various lines where molecules have been attached that excert specific interactions with the target. Here, we describe a colloidal gold-based immunochromatographic test strip for the simultaneous detection of ochratoxin (OTA) and zearalenone (ZEN) in corn and other cereals qualitatively and quantitatively.

Development of a Colloidal Gold-Based Immunochromatographic Strip for Rapid Detection of Klebsiella pneumoniae Serotypes K1 and K2.

In this study, a novel colloidal gold-based immunochromatographic strip (ICS) containing anti-Klebsiella pneumoniae capsular polysaccharide polyclonal antibodies was developed to specifically detect K. pneumoniae serotypes K1 and K2. Capsular polysaccharide K1 and K2 antigens were first used to produce polyclonal anti-K1 and anti-K2 antibodies. Reference strains with different serotypes, nontypeable K. pneumoniae strains, and other bacterial species were then used to assess the sensitivity and specificity of these test strips. The detection limit was found to be 105 CFU, and the ICSs were stable for 6 months when stored at room temperature. No false-positive or false-negative results were observed, and equivalent results were obtained compared to those of more conventional test methods, such as PCR or serum agglutination. In conclusion, the ICS developed here requires no technical expertise and allows for the specific, rapid, and simultaneous detection of K. pneumoniae serotypes K1 and K2.

Immunochromatographic strip assay for detection of bioactive Ganoderma triterpenoid, ganoderic acid A in Ganoderma lingzhi

Ganoderic acid A (GAA) is one of the major Ganoderma triterpenes produced by medicinal mushroom belonging to the genus Ganoderma (Ganodermataceae). Due to its interesting pharmacological activities, Ganoderma species have been traditionally used in China for the treatment of various diseases. Herein, we developed a colloidal gold-based immunochromatographic strip assay (ICA) for the rapid detection of GAA using highly specific monoclonal antibody against GAA (MAb 12A) conjugated with gold nanoparticles. Using the developed ICA, the detection of GAA can be completed within 15min after dipping the test strip into an analyte solution with the limit of detection (LOD) for GAA of ~500ng/mL. In addition, this system makes it possible to perform a semi-quantitative analysis of GAA in Ganoderma lingzhi, where high reliability was evaluated by enzyme-linked immunosorbent assay (ELISA). The newly developed ICA can potentially be applied to the standardization of Ganoderma using GAA as an index because GAA is major triterpenoid present much in the mushroom.

Development of a Colloidal Gold-based Immunochromatographic Test Strip for Detection of Cetacean Myoglobin

This protocol describes the development of a colloidal gold immunochromatographic test strip based on the sandwich format that can be used to differentiate the myoglobin (Mb) of cetaceans from that of seals and other animals. The strip provides rapid and on-the-spot screening for cetacean meat, thereby restraining its illegal trade and consumption. Two monoclonal antibodies (mAbs) with reactivity toward the Mb of cetaceans were developed. The amino acid sequences of Mb antigenic reactive regions from various animals were analyzed in order to design two synthetic peptides (a general peptide and a specific peptide) and thereafter raise the mAbs (subclass IgG1). The mAbs were selected from hybridomas screened by indirect ELISA, western blot and dot blot. CGF5H9 was specific to the Mbs of rabbits, dogs, pigs, cows, goats, and cetaceans while it showed weak to no affinity to the Mbs of chickens, tuna and seals. CSF1H13 can bind seals and cetaceans with strong affinity but showed no affinity to other animals. Cetacean samples from four families (Balaenopteridae, Delphinidae, Phocoenidae and Kogiidae) were used in this study, and the results indicated that these two mAbs have broad binding ability to Mbs from different cetaceans. These mAbs were applied on a sandwich-type colloidal gold immunochromatographic test strip. CGF5H9, which recognizes many species, was colloid gold-labeled and used as the detection antibody. CSF1H13, which was coated on the test zone, detected the presence of cetacean and seal Mbs. Muscle samples from tuna, chicken, seal, five species of terrestrial mammals and 15 species of cetaceans were tested in triplicate. All cetacean samples showed positive results and all the other samples showed negative results.

Development of a Colloidal Gold-based Immunochromatographic Test Strip for Detection of Cetacean Myoglobin.

This protocol describes the development of a colloidal gold immunochromatographic test strip based on the sandwich format that can be used to differentiate the myoglobin (Mb) of cetaceans from that of seals and other animals. The strip provides rapid and on-the-spot screening for cetacean meat, thereby restraining its illegal trade and consumption. Two monoclonal antibodies (mAbs) with reactivity toward the Mb of cetaceans were developed. The amino acid sequences of Mb antigenic reactive regions from various animals were analyzed in order to design two synthetic peptides (a general peptide and a specific peptide) and thereafter raise the mAbs (subclass IgG1). The mAbs were selected from hybridomas screened by indirect ELISA, western blot and dot blot. CGF5H9 was specific to the Mbs of rabbits, dogs, pigs, cows, goats, and cetaceans while it showed weak to no affinity to the Mbs of chickens, tuna and seals. CSF1H13 can bind seals and cetaceans with strong affinity but showed no affinity to other animals. Cetacean samples from four families (Balaenopteridae, Delphinidae, Phocoenidae and Kogiidae) were used in this study, and the results indicated that these two mAbs have broad binding ability to Mbs from different cetaceans. These mAbs were applied on a sandwich-type colloidal gold immunochromatographic test strip. CGF5H9, which recognizes many species, was colloid gold-labeled and used as the detection antibody. CSF1H13, which was coated on the test zone, detected the presence of cetacean and seal Mbs. Muscle samples from tuna, chicken, seal, five species of terrestrial mammals and 15 species of cetaceans were tested in triplicate. All cetacean samples showed positive results and all the other samples showed negative results.

Development of an immunochromatographic test strip for simultaneous qualitative and quantitative detection of ochratoxin A and zearalenone in cereal.

Ochratoxin A (OTA) and zearalenone (ZEN) are natural products of filamentous fungi that are harmful to humans and animals exposed to them even in extremely low concentration. The immunochromatographic test strip has become a popular diagnostic tool for detecting analytes. Its major advantages are that results can be obtained within 5-10 min, all needed reagents are included in the strip and it can be used to detect OTA and ZEN contamination in spots. In this study a colloidal gold-based immunochromatographic test strip of competitive format was developed for the rapid simultaneous qualitative and quantitative detection of OTA and ZEN in corn and other cereals. The test strip results with the naked eye showed that the sensitivities were 6 µg kg(-1) OTA and 20 µg kg(-1) ZEN in cereal, while those with a TSR3000 membrane strip reader showed that the IC50 values of OTA and ZEN were 1.7905 and 4.3514 ng mL(-1) and the lower detection limit (LDL) values were 0.7697 and 1.2000 µg kg(-1) respectively. These results were confirmed by high-performance liquid chromatography. The immunochromatographic test strip developed in this study could be used for the rapid simultaneous, qualitative and quantitative screening of OTA and ZEN in corn samples. © 2015 Society of Chemical Industry.

Development of an immunochromatographic strip for the serodiagnosis of Theileria infection in sheep.

BackgroundTheileria uilenbergi and T. luwenshuni are tick-borne protozoan parasites, transmitted by Haemaphysalis qinghaiensis and H. longicornis, respectively. They are the main causative agents of theileriosis in small ruminants in China. The disease has resulted in severe economic losses and hindered the development of sheep and goat husbandry industry in the endemic regions.MethodsIn this study, a colloidal gold-based immunochromatographic strip (ICS) was developed for the detection of T. uilenbergi and/or T. luwenshuni infections. A recombinant T. uilenbergi immunodominant protein (rTuIP) was used as antigen for the ICS. The nitrocellulose membrane was incubated with rTuIP on the test (T) line and anti-rTuIP antiserum on the control (C) line, respectively. The rTuIP conjugated to colloidal gold particles was used as the detection system for visualization of the lines. Then the sample pad, conjugate pad, nitrocellulose membrane and absorbent pad were assembled onto a backing plate in the appropriate order.ResultsThe ICS was able to detect antibodies in the sera of animals experimentally infected with T. uilenbergi from 14 to 85 days. It also reacted with the serum from T. luwenshuni infected sheep. However, there was no cross-reactivity with sera from animals infected with Babesia motasi and Anaplasma ovis. Comparison of the ICS with the rTuIP antigen based indirect enzyme-linked immunosorbent assays (ELISA) using test field samples showed good correlations with 93.1 % (81/87) sensitivity and 100 % (40/40) specificity, respectively, with an almost perfect agreement (Kappa = 0.895, P < 0.01).ConclusionAn immunochromatographic strip test based on a recombinant T. uilenbergi immunodominant protein (rTuIP) was developed. This is a rapid test (approximately 15 min to completion) for the detection of T. uilenbergi and/or T. luwenshuni infection that is easy to perform and; delivers results that are visible to the naked eye.