Preparation of colloidal gold immunochromatography strip for detection of methamidophos residue

Methamidophos (Met) is a broad spectrum organophosphorus insecticide and acaricide.Even a trace of its residue is harmful to humans and many animals.In this study,the synthesis and identification of colloidal gold particles and antibody-colloidal gold conjugates were performed,and the preparation of colloidal gold immunochromatography strip was conducted for detection of Met residue.The size of colloidal gold particles was checked using a transmission electron microscope (TEM).The formation of antibody-coll...

A New Method for Ultra-sensitive P24 Antigen Assay Based on Near-infrared Fluorescent Microsphere Immunochromatography

Objective To develop a rapid,highly sensitive quantitative method for detecting P24 antigen based on near-infrared fluorescent microsphere immunochromatography.Methods First,we prepared a lateral flow assay test strip,and labeled the detection antibody using a fluorescent microsphere.Second,we optimized the antibody labeling conditions.Third,we optimized the detection conditions.Fourth,we created a working curve.Fifth,we conducted a methodological assessment of the established fluorescent microsphere immunochromatography method.Sixty-six clinical samples were tested,and we compared the established fluorescent microsphere immunochromatography with the quantitative ELISA method.Results According to the working curve,the detection limit of the method is 3.4 pg/mL,and the detection range is 3.4 pg/mL to 10 ng/mL.The average intra-assay recovery was 99.6%,and the Coefficient of Variation(CV)was 5.4%–8.6%;the average inter-assay recovery was 97.3%,and the CV was 8.5%–11%.The detection rate of fluorescent microsphere immunochromatography was higher than ELISA method,and had a good correlation with ELISA.Conclusion The P24 antigen quantitative detection method based on near-infrared fluorescent microsphere immunochromatography has the advantages of rapid detection,high sensitivity,and wide detection range;thus,it is suitable for early clinical diagnosis and continuous monitoring of AIDS.

Study on Colloidal Gold Immunochromatography Assay for Rapid Detection of Spectinomycin

[Objectives] This study was conducted to establish a rapid detection method for spectinomycin in pork,chicken,fish,shrimp flesh and water.[Methods]A test strip for rapid detection of spectinomycin in milk was developed by colloidal gold immunochromatography assay. [Results]The test strip had a detection limit of 50 μg/kg to milk with a detection time of 15 min,and the false positive rate and false negative rate were both 0. [Conclusions]The method is accurate,simple,reliable and convenient,and is suitable for rapid on-site spectinomycin detection.

A novel colloidal gold immunochromatography assay strip for the diagnosis of schistosomiasis japonica in domestic animals

Background:Schistosomiasis remains a major public health concern in China and an epidemiological survey has revealed that schistosome-infected bovines and goats are the main transmission sources for the disease.Therefore,development of a sensitive technique for the diagnosis of schistosomiasis in domestic animals is necessary.Method:A novel colloidal gold immunochromatography assay(GICA)strip was developed for detecting Schistosoma japonicum in domestic animals.The colloidal gold was conjugated with recombinant streptococcal protein G(rSPG).As the test and control lines,the schistosome soluble egg antigen and rSPG,respectively,were blotted on nitrocellulose membrane.Results:The lowest detectable serum dilution was 1∶640 for schistosome-infected buffaloes.The cross-reaction rate of GICA was 14.29%with Paramphistomum sp.in buffaloes,16.67%with Haemonchus sp.in goats,and 33.33%with Orientobilharzia sp.in goats.These results were slightly lower and similar to those obtained through ELISA.Moreover,the strips for detecting S.japonicum in mice,rabbits,buffaloes,and goats showed high sensitivity(100.00%,100.00%,100.00%,and 100.00%,respectively)and specificity(100.00%,100.00%,94.23%,and 88.64%,respectively).And the sensitivity or specificity of the GICA strips did not present any significant differences after storage for 12 months at room temperature.When compared with ELISA,the GICA strips exhibited similar sensitivity and specificity in the diagnosis of schistosomiasis in mice,rabbits,buffaloes,and goats.Besides,only 5μl of serum are required for the test and the detection can be completed within 5 min.Conclusion:This study is the first to develop a GICA strip using gold-rSPG conjugate for the diagnosing of schistosomiasis in domestic animals,and preliminary results showed that the developed strip may be suitable for large-scale screening of schistosomiasis in endemic areas.

Stool antigen tests for the management of Helicobacter pylori infection

Stool antigen tests(SATs)are noninvasive diagnostic modules for Helicobacter pylori(H.pylori)infection.Two types of SATs exist for the diagnosis of H.pylori infection,one based on enzyme immunoassay(EIA)and another on immunochromatography(ICA).SATs do not require expensive chemical agents or specified equipment;hence,they are less expensive compared with the urea breath test.Both European and Japanese guidelines have shown that EIA-based SATs using monoclonal antibodies are useful for primary diagnosis as well as for the assessment of eradication therapy.ICA-based tests do not require particular equipment and are therefore useful in developing countries.SATs are also useful for the diagnosis of H.pylori infection in children and post gastric surgery patients.SATs performed via EIA can assess H.pylori infection in a large number of subjects,almost as well as serology.Thus,SATs would be useful or detecting current infection in such a survey to identify and eradicate H.pylori infection.The accuracy of SATs is lower when the stool samples are unformed or watery,because H.pylorispecific antigens in the stool samples are diluted.Temperature and the interval between stool sample collection and measurement also affect the results of SATs.The choice of test kit depends on the sensitivity and specificity in each region and the circumstances of each patient.

Machine Learning Approach to Enhance the Performance of MNP?Labeled Lateral Flow Immunoassay

The use of magnetic nanoparticle(MNP)-labeled immunochromatography test strips(ICTSs) is very important for point-ofcare testing(POCT). However, common diagnostic methods cannot accurately analyze the weak magnetic signal from ICTSs, limiting the applications of POCT. In this study, an ultrasensitive multiplex biosensor was designed to overcome the limitations of capturing and normalization of the weak magnetic signal from MNPs on ICTSs. A machine learning model for sandwich assays was constructed and used to classify weakly positive and negative samples, which significantly enhanced the specificity and sensitivity. The potential clinical application was evaluated by detecting 50 human chorionic gonadotropin(HCG) samples and 59 myocardial infarction serum samples. The quantitative range for HCG was 1–1000 mIU mL^(-1) and the ideal detection limit was 0.014 mIU mL^(-1), which was well below the clinical threshold. Quantitative detection results of multiplex cardiac markers showed good linear correlations with standard values. The proposed multiplex assay can be readily adapted for identifying other biomolecules and also be used in other applications such as environmental monitoring, food analysis, and national security.

Fast and Accurate Identification of <i>M. tuberculosis</i>Complex Using an Immunochromatographic MPT64 Antigen Detection Test

Background: A new rapid Immunochromatographic test (ICT) kit (MPT64 TB Ag Kit) for detection of MPT64 Antigen in M. tuberculosis (MTB) isolates used for rapid identification of MTB isolates developed by SD (Standard Diagnostics) Bio line, South Korea was evaluated. The ICT is a rapid, reliable and cheaper method that can be used instead of conventional biochemical tests for confirming MTB in culture isolates in resource limited laboratories. The study also evaluated the ability of ICT to detect MPT64-Antigen before the micro MGIT could signal positive. Material/Methods: A total of 450 sputum samples of individual patients were used for the study. 152 isolates of Mycobacteria were recovered from solid and liquid media. These strains were tested for the detection of MPT64-antigen. H37Rv strain was served as the positive reference control and also used for early detection of Antigen experiment. Findings: The development of bands on both test and sample region when H37Rv strain was tested were seen (MPT64 antigen positive). When 138 MTB isolates were tested, it showed a similar banding pattern indicating 100% sensitivity. MPT64 band formation was not detected in any of the 14 isolates indicating 100% specificity. Both PPV & NPV were 100%. All the isolates negative for MPT64 Ag were confirmed as MOTT by conventional bio-chemical PNBA. The H37Rv strain showed a faint band from the 2nd day onwards from inoculation till 3rd day in the earlier Antigen detection experiment. Conclusion: Rapid identification of MTB culture isolate is a pressing need for diagnosis and proceeding to perform drug susceptibility testing. MPT64 TB Ag detection ICT kit is a rapid, reliable method, good substitute for molecular identification methods, and conventional biochemical test which is time-consuming and technically demanding. The early detection of Antigen can be used as an effective tool in diagnosis.

Study on Rapid Detection of Tetracyclines,Fluoroquinolones and Sulfonamides in Milk

Aiming at the market demand for rapid detection of tetracyclines,fluoroquinolones and sulfonamides in milk,a golloidal gold immunochromatography test strip for simultaneous detection of tetracyclines,fluoroquinolones and sulfonamides in milk was prepared based on the principle of competitive inhibition immunochromatography. The performance indicators of the test strip were verified. The results showed that the test strip can simultaneously detect 4 tetracyclines,13 fluoroquinolones and 13 sulfonamides,and the detection limits all can meet the national residue limits; the tests strip exhibited false positive rate≤5% and false negative rate = 0; and no cross-reaction with other drugs was commonly found in milk,indicating good specificity. The method is simple,rapid,and has low cost and easy popularization. It provides a means for realizing on-site rapid detection and is of important practical significance to guarantee of safety of milk and dairy products in China.

The Incidence of Rotavirus and Adenovirus Infections among Children with Diarrhea in Sulaimani Province, Iraq

Human rotavirus and adenovirus infections are major causes of acute outbreaks and sporadic cases of gastroenteritis, occurring primarily among children less than 5 years of age. Little is known about the epidemiology of rotavirus and enteric adenovirus infections in Sulaimani and Iraq. The aim of this study was to determine the incidence and clinical significance of rotavirus and enteric adenovirus gastroenteritis and also to determine possible risk factors for rotavirus and adenovirus gastroenteritis using new simple rapid screening test (VIKIA ROTA AND ADENO). This is a qualitative test based on the immunochromatography technique. In the study, one hundred children less than 5 years of age with acute gastroenteritis admitted to Sulaimani Paediatric Hospital were studied. Rotavirus was identified in 22% of the children, adenovirus was identified in 3% of the children, and mixed rotavirus and adenovirus was identified in 2% of the children. All positive cases were younger than 2 years of age. The findings show that rotavirus is most commonly detected. However, there were no significant associations between rotavirus and adenovirus and gender, type of feeding, geographical distribution, the source of drinking water, and the past history of admission to hospital.

Preliminary Study on Immuno-gold Chromatography Assay for Rapid Detection of Vibrio parahaemolyticus

[ Objective] Vibrio parahaemolyticus was chosen as the material to make rabbit anti- V. parahaemolyticus polyclonal antibody, the immuno-gold chromatography assay （IGCA） was used to develop immunogold labeling test strips for detecting V. parahaemolyticus. [Methods] Rabbit anti-V, parahaemolyticus IgG-2 was used to wrap nitrocellulose membrane detection line, goat anti-rabbit IgG was used to wrap control line, and im- muno-gold was used to label rabbit anti-V, parahaemolyticus IgG-1 to establish the immuno-gold chromatography assay （IGCA） for detection of V. parahaemolyticus. [Results] Test strip detection lines of positive results along with the control line were all red, test results could be got in only five to fifteen minutes, the minimum detectable amount of this method to V. parahaemolyticus was 3.60 x 104 cfu/ml, and cross reactions wouldnt occur when detecting common intestinal bacteria like Vibrio alginolyticus, vibrio cholerae, Vibrio damsela, Vibrio metschnikovii, Citrobacter freundii and salmonella etc. The test results were undifferentiated when the test strips were stored at 4 ~C for six months, stored at room temperature for three months or stored at 37 ~C for one month. [ Condusionl The established test strip assay was simple and rapid during operation, with high sensitivity, strong specificity and good stability, the test results were easy to be observed and judged, and this assay was very suitable for grassroots breeding department application.

An Immunochromatographic Strip Test for Rapid Detection of Antibodies against Porcine Circovirus Type 2

Porcine cimovirus type 2 (PCV2) infection causes huge economic losses, but no serological method is available for batch detection of field samples. The aim of the study was to develop a method for large-batch detection of PCV2 infection. Colloidal gold-labeled staphylococcal protein A (SPA) was sprayed on glass fibers to prepare a conjugate pad. The recombinant ORF2 protein of PCV2 was blotted on the test line of the nitrocellulose (NC) membrane, and pig IgG was streaked on the control line of the NC membrane. Then, the immunochromatographic strip was used for detection of antibodies against PCV2. The results show that the strip test was simple and the results could be determined within 10 min with naked eyes. The test strip was highly specific for pig serum against PCV2 and no cross-reaction with pig serum against other pathogens was observed. The test strip had close similarity with ELISA. Storage at RT for 6 months did not affect the specificity and sensitivity obviously. A total of 324 clinical pig sera were detected by both ELISA and the developed test strip, and the coincidence was 98.77%. Therefore, the developed immunochromatographic strip is specific, sensitive, stable, fast and simple, and it is suitable for on-site detection of antibodies against PCV2.

Hepatitis C Status in Karachi, Pakistan, a Five-Year Survey at Civil Hospital, Karachi, Pakistan

Hepatitis C virus is associated with Hepatitis C which can be acute or chronic. Acute hepatitis C is usually short term illness that can lead to chronic infection, while chronic hepatitis C is a chronic liver disease leading to cirrhosis, hepatocellular carcinoma and even death. The mode of transmission is mainly parenteral and vertical. Hepatitis C is the leading cause of morbidity and mortality and a serious public health problem, worldwide as well as in Pakistan. There is no vaccine available for hepatitis C so the only way to prevent hepatitis C is by changing behaviors which can spread the disease. Pakistan is also facing a huge burden of this disease. In Pakistan, the frequency of hepatitis C virus infection ranges from 8% - 15% in the general population with variations in different parts of the country. The general method of detecting infection with HCV is to observe the presence of antibodies to the virus. The aim of this study was to determine frequency of hepatitis C among general population and to see the increasing frequency in our part. Selection of Patients: All patients visiting outpatient clinics, Hepatitis clinic and indoor patients at Civil Hospital Karachi, Pakistan from January 1, 2010 to December 31, 2014 were screened for anti HCV. Methods: Blood samples were collected from the patients in Red Topped vaccutainers and allowed to clot, then centrifuged and serum was screened for anti HCV by Immunochromatography (ICT) method. Results: A total of 156,990 patients were screened for anti-HCV, 23,511 (14.98%) patients were reported positive. During the year 2010, total 11,058 patients were screened for anti HCV, 1791 (16%) were positive, while in the year 2011 total 26,921 were screened and 3908 (14.5%) were positive, during the year 2012, total 23,321 were screened, 3036 (14%) were positive, in the year 2013 we screened total 40,483 patients 6070 (15%) were positive, while in 2014, total 55,484 patients were screened and 8584 were positive (15.5%). The most affected age group is between 21 - 40 years and males are affected more than females. Conclusion: Hepatitis C in increasing in our population affecting the young generation between the age of 21 - 40 years and male population is affected more than females so a mass screening is needed to know the exact position of the disease as well as awareness programs should be started using media.

Immune fluorescence test strips based on quantum dots for rapid and quantitative detection of carcino-embryonic antigen

At present,many researchers focused on the point-of-care testing（POCT）,a method of disease markers detection without large-scale instruments and specialized persons.However,most POCT diagnostic methods were suffered from poor detection sensitivity or inefficiency in quantitative detection.Herein,we developed a newly QD-immune fluorescence test strips（QD-IFTS） based on quantum dots（QDs） as the fluorescence nanocarrier to prepare the immune fluorescence probes in the classical immunochromatography detection system for sensing carcino-embryonic antigen（CEA）,a kind of glycoprotein produced by intestinal tissue and a broad spectrum of tumor marker for cancer diagnosis.And we designed a homemade strips fluorescence reader for detection of fluorescence intensity of QDs on the QD-IFTS.Under the optimized reaction conditions,chromatographic time of the newly QD-IFTS was only25 min,sample volume of the newly QD-IFTS was only 40 m L and the LOD of the newly QD-IFTS was 0.72 ng/m L.In addition,the efficiency and robustness of the newly QD-IFTS were confirmed by successfully application in 300 clinical serum samples,and the results revealed great potential in clinical POCT of other biomarkers.

IMMUNOCHEMICAL IDENTIFICATION OF SPECTRINS ON THE PLASMA MEMBRANE OF LEAF CELLS OF Vicia faba

It was well established recently that a membrane skeleton was present on the plasma membrane of erythrocytes and nonerythrocytes of animals. The membrane skeleton plays an important role in the maintenance of cell shape, regulation of cell motility and stabilizing the lipid bilayer. In the previous paper we compared the membrane proteins of the plasma membrane of leaf cells of Vicia faba with the membrane skeleton pro-