Enzyme Immunoassay For Detection Research Articles

Serosurveillance for Measles and Rubella.

Measles and rubella remain global health threats, despite the availability of safe and effective vaccines. Estimates of population immunity are crucial for achieving elimination goals and assessing the impact of vaccination programs, yet conducting well-designed serosurveys can be challenging, especially in resource-limited settings. In this review, we provide a comprehensive assessment of 130 measles and rubella studies published from January 2014 to January 2024. Methodologies and design aspects of serosurveys varied greatly, including sample size, assay type, and population demographics. Most studies utilized enzyme immunoassays for IgG detection. Sample sizes showed diverse sampling methods but favored convenience sampling despite its limitations. Studies spanned 59 countries, predominantly including adults, and revealed disparities in seroprevalence across demographics, regions, and notably among migrants and women. Age-related declines in antibodies were observed, particularly among infants, and correlations between vaccination status and seropositivity varied. We conclude with an outlook on measles and rubella serosurveillance, emphasizing the need for proper survey design and the advantages of standardized, multiplex serology assays.

Plasmonic enzyme immunoassay via nanobody-driven controllable aggregation of gold nanoparticles for detection of ochratoxin A in pepper

Ochratoxin A (OTA) in food poses a serious challenge to public health. Herein, using the nanobody-driven controllable aggregation of gold nanoparticles (AuNPs) in a glucose oxidase-tyramine-horseradish peroxidase (GOx-TYR-HRP) system, we propose a direct competitive plasmonic enzyme immunoassay (dc-PEIA) for OTA detection. The OTA-GOx conjugate catalyzes glucose to produce hydrogen peroxide (H2O2), and then HRP catalyzes H2O2 to generate hydroxyl radical which induces the crosslink of TYR. Crosslinked TYR leads to aggregation of AuNPs through strong electrostatic interactions, which is tunable based on the competition of OTA-GOx and free OTA for binding the immobilized nanobody. The optimized dc-PEIA achieves an instrumental limit of detection (LOD) of 0.275 ng/mL and a visual LOD of 1.56 ng/mL. It exhibits good selectivity for OTA and accuracy in the analysis of pepper samples, with the confirmation of high-performance liquid chromatography. Overall, the dc-PEIA is demonstrated as a useful tool for detecting OTA in food.

Verification of the precision of enzyme immunoassay detection of HBsAg in «Vectohep B — HBs-antigen» reagent kits

Verification of the precision of enzyme immunoassay detection of HBsAg in «Vectohep B — HBs-antigen» reagent kits

Semi-quantitative graphene chemiresistor enzyme immunoassay for simple and sensitive antigen detection

Graphene is an ideal platform for sensing biomolecule charges due to its high electron/hole mobilities and 2D nature. However, when directly detecting biomolecules on graphene surfaces, it is important to consider their charge magnitude and the impact of Debye screening, which limits the detectable biomolecule types. Therefore, we propose a graphene chemiresistor enzyme immunoassay that allows for the detection of all protein types, regardless of charge or size. In this study, we immobilized antibodies on rigid glass surfaces rather than graphene surfaces to simplify the washing process and reduce noise caused by solution flow on graphene. As an example, we used influenza virus nucleoprotein as the antigen and urease as the enzyme. By measuring changes in graphene's electrical conductivity resulting from ammonium ions generated by enzymatic reactions, we confirmed antigen presence. The responsiveness of the graphene chemiresistor varied depending on antigen concentration, successfully detecting 100 pM antigens in this experiment. Our semi-quantitative method offers a simple and highly sensitive electrical detection of antigens.

Evaluation of an enzyme immunoassay and immunodiffusion for detection of anti-Histoplasma antibodies in serum from cats and dogs.

Histoplasma antigen and anti-Histoplasma antibody detection are used to support the diagnosis of histoplasmosis. There is a paucity of published data on antibody assays. Our primary hypothesis was that anti-Histoplasma immunoglobulin G (IgG) antibody detection using enzyme immunoassay (EIA) will be more sensitive as compared to immunodiffusion (ID). Thirty-seven cats and 22 dogs with proven or probable histoplasmosis; 157 negative control animals. Residual stored sera were tested for anti-Histoplasma antibodies using EIA and ID. Results of urine antigen EIA were reviewed retrospectively. Diagnostic sensitivity was calculated for all three assays and compared between immunoglobulin G (IgG) EIA and ID. The diagnostic sensitivity of urine antigen EIA and IgG EIA, interpreted in parallel, was reported. Sensitivity of IgG EIA was 30/37 (81.1%; 95% confidence interval [CI], 68.5%-93.4%) in cats and 17/22 (77.3%; 95% CI, 59.8%-94.8%) in dogs. Diagnostic sensitivity of ID was 0/37 (0%; 95% CI, 0%-9.5%) in cats and 3/22 (13.6%; 95% CI, 0%-28.0%) in dogs. Immunoglobulin G EIA was positive in all animals (2 cats and 2 dogs) with histoplasmosis but without detectable antigen in urine. Diagnostic specificity of IgG EIA was 18/19 (94.7%; 95% CI, 74.0%-99.9%) in cats and 128/138 (92.8%; 95% CI, 87.1%-96.5%) in dogs. Antibody detection by EIA can be used to support the diagnosis of histoplasmosis in cats and dogs. Immunodiffusion has an unacceptably low diagnostic sensitivity and is not recommended.

Rapid determination of SARS-CoV-2 antibody neutralization titer using Bio-Rad Bio-Plex correlates strongly with pseudovirus-determined neutralization titer

Accurate and rapid evaluation of SARS-CoV-2 half-maximal neutralizing antibody (nAb) titer (NT50) is an important research tool for measuring nAb responses after prophylaxis or therapeutics for COVID-19 prevention and management. Compared with ACE2-competitive enzyme immunoassays for nAb detection, pseudovirus assays remain low-throughput and labor intensive. A novel application of the Bio-Rad Bio-Plex Pro Human SARS-CoV-2 D614G S1 Variant nAb Assay was used to determine NT50 from COVID-19-vaccinated individuals and showed strong correlation to a laboratory-developed SARS-CoV-2 pseudovirus nAb assay. The Bio-Plex nAb assay could provide a rapid, high-throughput, culture-free method for NT50 determination in sera.

Development of indirect competitive ELISA and CLEIA for quantitative analysis of melatonin in health products.

Melatonin (MT), as a hormone regulating the rhythm of sleep, is widely used in health products. However, illicit and excessive use of MT might cause undesirable effects. Therefore, it is essential to establish highly sensitive and specific rapid methods for MT analysis in health products. In this study, we established indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) and indirect competitive chemiluminescent enzyme immunoassay (ic-CLEIA) for sensitive and selective detection of MT in health products. Under optimal conditions, half-maximum inhibitory concentration (IC50 value) and limit of detection (LOD, IC10 value) for MT by ic-ELISA were 0.25 ng mL-1 and 0.03 ng mL-1 respectively, while the IC50 and LOD of ic-CLEIA were lower at 0.17 ng mL-1 and 0.03 ng mL-1 respectively. Three MT-free Chinese patent medicines were spiked with MT and the recovery rates ranged from 71.89% to 117% (ic-ELISA) and 83.66% to 107.17% (ic-CLEIA). The level of MT in six MT-containing health products was assessed in parallel using the developed methods and HPLC. Both ic-ELISA and ic-CLEIA showed good consistencies (R2 = 0.999 and 0.993, respectively) with HPLC, indicating that the two methods developed were sensitive, fast, and reliable for application in MT analysis.

Mst1 silencing alleviates hypertensive myocardial injury associated with the augmentation of microvascular endothelial cell autophagy.

The activation of mammalian ste20‑like kinase1 (Mst1) is a crucial event in cardiac disease development. The inhibition of Mst1 has been recently suggested as a potential therapeutic strategy for the treatment of diabetic cardiomyopathy. However, whether silencing Mst1 also protects against hypertensive (HP) myocardial injury, or the mechanisms through which this protection is conferred are not yet fully understood. The present study aimed to explore the role of Mst1 in HP myocardial injury using invivo and invitro hypertension (HP) models. Angiotensin II (Ang II) was used to establish HP mouse and cardiac microvascular endothelial cell (CMEC) models. CRISPR/adenovirus vector transfection was used to silence Mst1 in these models. Using echocardiography, hematoxylin and eosin staining, Masson's trichrome staining, the enzyme‑linked immunosorbent assay detection of inflammatory factors, the enzyme immunoassay detection of oxidative stress markers, terminal deoxynucleotidyl transferase dUTP nick‑end labeling staining, scanning electron microscopy, transmission electron microscopy, as well as immunofluorescence and western blot analysis of the autophagy markers, p62, microtubule‑associated proteins1A/1B light chain 3B and Beclin‑1, it was found that AngII induced HP myocardial injury with impaired cardiac function, increased the expression of inflammatory factors, and elevated oxidative stress in mice. In addition, it was found that Ang II reduced autophagy, enhanced apoptosis, and disrupted endothelial integrity and mitochondrial membrane potential in cultured CMECs. The silencing of Mst1 in both invivo and invitro HP models attenuated the HP myocardial injury. On the whole, these findings suggest that Mst1 is a key contributor to HP myocardial injury through the regulation of cardiomyocyte autophagy.

A2A R mediated modulation in IP3 levels altering the [Ca2+]i through cAMP-dependent PKA signalling pathway

Stimulation of A2A receptors (A2A R) coupled to Gs/olf protein activates Adenylyl cyclase (AC) leading to the release of cAMP which activates the cAMP-dependent PKA phosphorylation. The possible role of A2A R in the modulation of free cytosolic Ca2+ concentration ([Ca2+]i) involving IP3, cAMP and PKA was investigated in HEK 293-A2A R. The levels of IP3 and cAMP were observed by enzyme immunoassay detection method and [Ca2+]i using Fluo-4 AM. Moreover, cAMP-dependent PKA was determined using the PKA Colorimetric Activity Kit. We observed that the cells pre-treated with A2A R agonist NECA showed increased levels of cAMP, PKA, IP3 and [Ca2+]i levels. However, the reverse effect was observed with A2A R antagonists (ZM241385 and caffeine). Blocking the Gαq/PLC/DAG/IP3 pathway with neomycin, a PLC inhibitor did not affect the modulation of IP3 and [Ca2+]i levels in HEK 293-A2A R cells. To investigate the Gαi/AC/cAMP/PKA, HEK 293-A2A R cells pre-treated with pertussis toxin followed by forskolin in the presence of A2A R agonist (NECA) showed no effect on cAMP levels. Further, Gαs/AC/cAMP/PKA pathway was investigated to elucidate the role of cAMP-dependent PKA in IP3 mediated [Ca2+]i modulation. In the HEK 293-A2A R cells pre-treated with PKA inhibitor KT5720 and treated with NECA led to inhibit the IP3 and [Ca2+]i levels. The study distinctly demonstrated that A2A R modulates IP3 levels to release the [Ca2+]i via cAMP-dependent PKA. The role of A2A R mediated Gαs pathway inducing IP3 mediated [Ca2+]i release may open new avenues in the therapy of neurodegenerative disorder.

Serological diagnostic potential of the 38-72 kDa somatic antigen of Gastrothylax crumenifer in buffaloes using the Indirect Enzyme-Linked Immunosorbent Assay

Paramphistomosis, caused by digenetic trematodes of the superfamily Paramphistomoidea, causes heavy economic losses in terms of reduced fertility, milk and meat production in the livestock industry. In the present study, somatic and excretory secretory antigens isolated from 500 live Gastrothylax crumenifer were assessed for their diagnostic potential by using an antibody detection enzyme immunoassay. Prior to the enzyme immunoassay, the somatic and excretory/secretory (ES) antigens of G. crumenifer were subjected to SDS-PAGE and Western blot (WB) for the detection of immunogenic proteins. Indirect ELISA analysis was performed on sera from buffaloes naturally infected with G. crumenifer, with control sera of buffaloes infected with Gigantocotyle explanatum, Fasciola spp., Cotylophoron /Paramphistomum spp. The SDS-PAGE results of the somatic products of G. crumenifer identified proteins were between 10-123 kDa, showing a maximum abundance of 10, 15, 25-28, 36, 38-72, 95-123 kDa proteins. The most abundant proteins recorded in ES product were ≥95, 72, 55 and 40 kDa. The antigenic analysis of somatic proteins using WB revealed reactive polypeptides of a size between 55-70 kDa, while metabolic extracts did not show reactivity with naturally infected buffalo sera. The sensitivity and specificity of the ELISA test for 38-72 kDa somatic antigens were 85.71% and 89.74%, respectively. The cross-reactivity with other trematode sera was 16-20%. Antibodies were tested against the 38-72 kDa somatic antigens, and 19.69% (39/198) of buffaloes were found positive, while 12.1% (24/198) presented infection in the fecal/postmortem examination. The study confirmed that ELISA established for the 38-72 kDa somatic antigens of G. crumenifer had diagnostic potential against paramphistome infections.

Effect of Curcumin at Various Doses on the Pharmacokinetic Profile of Tacrolimus in Healthy Rabbits

The purpose of present study is to evaluate the effect of co-administration of curcumin (CUR) at various doses on the pharmacokinetic (PK) profile of tacrolimus (TAC), a CYP 3A4 substrate in healthy male rabbits. Healthy male rabbits (n=18) were employed in an in vivo, parallel-randomized study. Three groups of rabbits were selected and separated: The rabbits in the first group (control group) received 1 mg/kg TAC orally. Blood samples (1.5-2 mL) were drawn from rabbits' ear marginal veins at the following time frames: 15.0, 30.0, 45.0, 60.0, 90.0, 120.0, 150.0, 180.0 and 300 minutes after TAC administration post dosing and analyzed by using a TAC chemiluminescent enzyme immunoassay (CLIA) detection kit. In the second and third groups (test groups), rabbits received TAC (1mg/kg) at identical conditions as in the control group with volumes equivalent to (30 and 90 mg/kg/day) of prepared CUR suspension in normal saline for seven continuous days. Blood samples from the control group were obtained on the eighth day. Non-compartmental analysis was used to derive different PK parameters of TAC for the three groups. When CUR was co-administered at both concentrations, insignificant statistically small changes between the control and testing groups were found. Our results revealed that the differences for the three groups in PK parameters as Cmax, tmax, ke, AUC0-6 and AUC0-? were statistically insignificant (P>0.05). In conclusion, it has been found that CUR at the experimented doses does not affect the PK of TAC. Further confirmation of our findings is requiered before these results can be applied in patient care.

Comparison of a chemiluminescence immunoassay and an enzyme immunoassay for detection of IgM antibodies against measles, mumps, rubella, cytomegalovirus (CMV), Epstein Barr virus (EBV), and human herpes virus (HHV) -1 and -2 infections

PurposeOutbreaks of vaccine-preventable viral diseases have been increasingly reported globally over the past few years. The burden of congenital viral infections, their impact on physical and mental development and the resulting economic loss to the family and the community are also well known. IgM antibody detection has been convenient in the diagnosis of acute viral infections, particularly in settings with limited resources where molecular tests are not feasible. MethodsThis is a comparative study between a chemiluminescence immunoassay (Liaison, DiaSorin, Saluggia, Italy) and an enzyme linked immunosorbent assay (ELISA) (Euroimmun, Lubeck, Germany) for the detection of IgM antibody against measles, mumps, rubella, CMV, EBV and HHV-1 and -2 viruses using a total of 345 samples. Results are expressed as agreement using kappa statistics. ResultsIn this study, CLIA is perfectly comparable to ELISA for the detection of IgM antibodies against measles (0.86) and mumps (0.92) with a moderate agreement for rubella (0.52), CMV (0.57), EBV (0.50), and HHV-1 and -2 (0.47) assays. However, a PABAK (prevalence-adjusted bias-adjusted kappa) showed improved agreement for rubella (0.64), CMV (0.65), EBV (0.60), and HHV-1 and -2 (0.88) assays. ConclusionsIgM antibody assays (CLIA and ELISA) against measles and mumps virus can be comparably used depending on the laboratory setup, throughput and expertise.

Effect of Oral Administration of Valerian Extract at Different Doses on Pharmacokinetic Parameters of Carbamazepine in Rabbits

Carbamazepine (CBZ) is a narrow therapeutic index drug used in the treatment of trigeminal neuralgia and psychiatric disorders. Valerian (VAL) is a popular herbal product which should be prescribed to treat insomnia and anxiety. The study was designed to investigate the presence of significant pharmacokinetic (PK) interaction between Valerian (VAL) at different concentrations on Carbamazepine (CBZ) pharmacokinetic parameters in healthy male rabbits. In an in vivo, parallel-randomized controlled trial, the rabbits in three groups "first (control), second and third" were given oral doses of CBZ (50 mg/kg), for "second and third" groups (as test groups) rabbits were given (20 and 40 mg/kg/day) of the VAL respectively, as suspension in normal saline for eight consecutive days. On the eighth day, CBZ was co-administered an hour after adding the last dose of VAL suspension. Venous blood samples (1.0-1.5 mL) were obtained from rabbits' ears' marginal vein at predetermined different periods. The plasma of this blood separation was done using centrifugation and stored at -80°C, prior to analysis by using CBZ chemiluminescent enzyme immunoassay detection kit. Different PK parameters such as Cmax, tmax, t½, ke, AUC0-24 and AUC0-∞ were determined for the three groups, applying Statistical testing (ANOVA). The results showed statistical insignificant differences for all PK parameters among the three groups with (p˃0.05). The findings showed that VAL at both concentrations is not likely to interfere with PK parameters related to CBZ, Further confirmation in humans shoud be done before these findings are applied to patient care.

Seroprevalence and changing trend of dengue in a tertiary care hospital, Bhubaneswar, Odisha: Four-year retrospective study

Dengue virus is a single stranded positive sense RNA virus belonging to the genus flavivirus family flaviviridae. Dengue fever is a seasonal and emerging acute mosquito borne arbo-viral illness affecting tropical and sub-tropical countries. This illness ranges from a mild asymptomatic form to severe dengue hemorrhagic fever (DHF) with or without dengue shock syndrome (DSS). This study was conducted to know the seroprevalence and changing trend of Dengue virus in a tertiary care hospital, Bhubaneswar, Odisha, India. Over a period of four years from August 2016 to December 2019, a total of 5147 blood samples from clinically suspected dengue patients were received in department of Microbiology laboratory. Serum was separated and subjected to enzyme immunoassay for detection of both Non Structural (NS1) antigen and IgM antibody.During this study period, a total of 5147 blood samples were processed from suspected dengue cases, out of which 1314(25%) samples were found to be positive by different serological markers like NS1 Antigen (Ag), IgM antibody(Ab), or both NS1 Ag & IgM Ab. The overall seroprevalence rate was found to be 25%. In this study period of four years, the year-wise seroprevalence rate was found to be 12% (153) in the year 2016 and was 26% (350) in the year 2017 and was 40% (522) in the year 2018, and was 22% (289) in the year 2019. It clearly shows that there is an increase in the dengue cases.

Diagnostic potential of 36-55 kDa somatic antigens of Fasciola gigantica for bovine fasciolosis

Fascioliasis causes huge economic losses in livestock industry by reducing the growth, fertility rate, meat and milk yield. In the present study, somatic antigens isolated from Fasciola gigantica were assessed for the early detection of infection by developing antibody detection enzyme immunoassay. The somatic antigens extracted from adult F. gigantica worms was separated on SDS-PAGE and immunogenicity was determined by Western blot (WB). The results showed polypeptides bands between 15 to 95 kDa, and most prominent bands were 15, 28, 36, 38, 55, 72 and 95 kDa polypeptides. The WB revealed a single polypeptide of size between 36 to 55 kDa as antigenic. The sensitivity and specificity of ELISA test established for 36 to 55 kDa somatic antigens was 95.45% (95% CI: 77.16% to 99.88%) and 87.1% (95% CI: 70.17% to 96.37%), respectively. Kappa value revealed that the strength of agreement is considered to be very ‘good’. In house established ELISA was implemented in the field and 12.1% (31/256) large ruminants were found positive, while 4.68% (12/256) with postmortem examination. The fasciolosis did not show significant (P>0.05) association with host type, breed and age groups, while significant (P<0.01) association with host sexes. The result confirmed that in-house established ELISA test had good value for serodiagnosis of fasciolosis in cattle and buffaloes for large scale epidemiological studies.

Development and Validation of an Enzyme Immunoassay for Detection and Quantification of SARS-CoV-2 Salivary IgA and IgG.

Oral fluids offer a noninvasive sampling method for the detection of Abs. Quantification of IgA and IgG Abs in saliva allows studies of the mucosal and systemic immune response after natural infection or vaccination. We developed and validated an enzyme immunoassay (EIA) to detect and quantify salivary IgA and IgG Abs against the prefusion-stabilized form of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike protein expressed in suspension-adapted HEK-293 cells. Normalization against total Ab isotype was performed to account for specimen differences, such as collection time and sample volume. Saliva samples collected from 187 SARS-CoV-2 confirmed cases enrolled in 2 cohorts and 373 prepandemic saliva samples were tested. The sensitivity of both EIAs was high (IgA, 95.5%; IgG, 89.7%) without compromising specificity (IgA, 99%; IgG, 97%). No cross-reactivity with endemic coronaviruses was observed. The limit of detection for SARS-CoV-2 salivary IgA and IgG assays were 1.98 ng/ml and 0.30 ng/ml, respectively. Salivary IgA and IgG Abs were detected earlier in patients with mild COVID-19 symptoms than in severe cases. However, severe cases showed higher salivary Ab titers than those with a mild infection. Salivary IgA titers quickly decreased after 6 wk in mild cases but remained detectable until at least week 10 in severe cases. Salivary IgG titers remained high for all patients, regardless of disease severity. In conclusion, EIAs for both IgA and IgG had high specificity and sensitivity for the confirmation of current or recent SARS-CoV-2 infections and evaluation of the IgA and IgG immune response.

Tuning the surface enhanced Raman spectroscopy performance of Au core-Ag shell nanostructure for label-free highly sensitive detection of colorectal cancer Marker

Early detection is vital for the prevention and treatment of patients with colorectal cancer (CRC). However, existent methods (such as enzyme immunoassay, chemiluminescent immunoassay and radioimmunoassay) for early detection of CRC are time consuming, costly and complicated multistage processes, limiting their further applications. Here, a novel label-free and ultrasensitive detection method is developed for CRC protein marker NDKA by Cr3+ ions aggregating Au@Ag NPs (ICNPs) via surface-enhanced Raman spectroscopy (SERS) spectra for the first time. The Au@Ag NPs with size-controlled Ag shell thickness (1 nm, 3 nm, 5 nm, 7 nm and 9 nm Ag shells) are prepared as the SERS substrates and its Raman signals are Ag shells thickness-dependent sensitivity to proteins. By using Cr3+ ions as aggregating agents, the aggregating effect between the SERS substrates of Au@Ag ICNPs and protein analyte results in significantly enhanced and reproducible Raman signals owing to formation of lots of hotspots. Using this novel method, several label-free proteins, myoglobin (3 fg/mL), human serum albumin (30 fg/mL), lysozyme (30 ng/mL) and cytochrome C (30 ng/mL) were also detected at ultralow concentrations with good reproducibility and high sensitivity, demonstrating the potential applications in the fields of medical therapy and clinical diagnosis.

Pediatric Clostridioides difficile Infection: Diagnosis and Diagnostic Stewardship.

Although the pathogenesis of Clostridioides difficile infection (CDI) is complex and incompletely understood, it is believed that the elaboration of C. difficile toxins is necessary for disease. There are a variety of tests available for the detection of both the C. difficile organism and its toxins; however, each has limitations and the best application of these tests to the diagnosis of CDI in children remains uncertain. Nucleic acid amplification tests are unable to reliably discriminate between CDI and C. difficile colonization, while commercially available enzyme immunoassays for toxin detection lack sensitivity. An understanding of preanalytic factors, relevant patient features, and test performance characteristics is essential to the accurate diagnosis of CDI in children. Specific diagnostic stewardship strategies can also increase the likelihood that positive tests reflect disease rather than colonization. Ultimately, CDI remains a clinical diagnosis and clinical judgment is essential when interpreting test results, regardless of the methods used.

The laboratory test procedure to confirm rotavirus vaccine infection in severe complex immunodeficiency patients.

The rotavirus vaccine is a live vaccine, and there is a possibility of infection by the virus strain used in the vaccine. We investigated the process of determining whether an infection was caused by the vaccine strain in a severe complex immunodeficiency (SCID) patient with rotavirus infection. The patient was vaccinated with RotaTeq prior to being diagnosed with SCID. The testing process was conducted in the following order: confirming rotavirus infection, determining its genotype, and confirming the vaccine strain. Rotavirus infection was confirmed through enzyme immunoassay and VP6 gene detection. G1 and P[8] were identified by multiplex polymerase chain reaction for the genotype, and G3 was further identified using a single primer. By detecting the fingerprint gene (WC3) of RotaTeq, it was confirmed that the detected virus was the vaccine strain. Genotypes G1 and P[8] were identified, and the infection was suspected of having been caused by rotavirus G1P[8]. G1P[8] is the most commonly detected genotype worldwide and is not included in the recombinant strains used in vaccines. Therefore, the infection was confirmed to have been caused by the vaccine strain by analyzing the genetic relationship between VP4 and VP7. Rotavirus infection by the vaccine strain can be identified through genotyping and fingerprint gene detection. However, genetic linkage analysis will also help to identify vaccine strains.

Detection of Blastomyces dermatitidis Antigen in Urine Using a Commercially Available Quantitative Enzyme Immunoassay.

Laboratory diagnosis of blastomycosis relies on a combination of methods, including antigen detection. We assessed the performance of analyte-specific reagents from Gotham Biotech (Portland, ME) for quantitative detection of Blastomyces dermatitidis galactomannan (GM) in urine using an enzyme immunoassay (EIA) compared to the Blastomyces quantitative EIA from MiraVista Diagnostics (Indianapolis, IN). Residual urine from 232 unique patients previously tested by the MiraVista assay was evaluated using the Gotham EIA, which showed 97.4% (74/76), 100% (156/156), and 99.1% (230/232) positive, negative, and overall agreement, respectively. Correlation between the quantitative B. dermatitidis antigen levels by the Gotham and MiraVista EIAs was low (R2 = 0.20). Medical records were available for 36 of the 232 patients, among whom four had confirmed blastomycosis and both the Gotham and MiraVista EIAs were positive. Nine of these patients had histoplasmosis, and the Gotham and MiraVista EIAs yielded negative results in 44.4% (4/9) and 22.2% (2/9) of cases, respectively. Both assays were negative in the remaining 23 patients. After laboratory implementation of the Gotham EIA, chart reviews were performed on the first 50 unique patients (51 samples) tested by the assay in our hospital. Among these, 3/50 (6%) samples were positive by the Gotham EIA, including two samples from a patient with culture-confirmed blastomycosis and one from a patient with histoplasmosis (also positive by the MiraVista Blastomyces EIA). All remaining patients were negative by the Gotham EIA and had alternative diagnoses. Our findings show comparable performance between the Gotham and MiraVista quantitative EIAs for detection of B. dermatitidis GM in urine.