Establishment and performance analysis of a new multiplex detection method for influenza an and B virus antigen

BACKGROUND Influenza A and B virus detection is pivotal in epidemiological surveillance and disease management.Rapid and accurate diagnostic techniques are crucial for timely clinical intervention and outbreak prevention.Quantum dot-encoded microspheres have been widely used in immunodetection.The integration of quantum dot-encoded microspheres with flow cytometry is a well-established technique that enables rapid analysis.Thus,establishing a multiplex detection method for influenza A and B virus antigens based on flow cytometry quantum dot microspheres will help in disease diagnosis.AIM To establish a codetection method of influenza A and B virus antigens based on flow cytometry quantum dot-encoded microsphere technology,which forms the foundation for the assays of multiple respiratory virus biomarkers.METHODS Different quantum dot-encoded microspheres were used to couple the monoclonal antibodies against influenza A and B.The known influenza A and B antigens were detected both separately and simultaneously on a flow cytometer,and the detection conditions were optimized to establish the influenza A and B antigen codetection method,which was utilized for their detection in clinical samples.The results were compared with the fluorescence quantitative polymerase chain reaction(PCR)method to validate the clinical performance of this method.RESULTS The limits of detection of this method were 26.1 and 10.7 pg/mL for influenza A and B antigens,respectively,which both ranged from 15.6 to 250000 pg/mL.In the clinical sample evaluation,the proposed method well correlated with the fluorescent quantitative PCR method,with positive,negative,and overall compliance rates of 57.4%,100%,and 71.6%,respectively.CONCLUSION A multiplex assay for quantitative detection of influenza A and B virus antigens has been established,which is characterized by high sensitivity,good specificity,and a wide detection range and is promising for clinical applications.

The combined detection of carcinoembryonic antigen,carcinogenic antigen 125,and carcinogenic antigen 19-9 in colorectal cancer patients

BACKGROUND Hepatic metastases are common and difficult to treat after colorectal cancer(CRC)surgery.The predictive value of carcinoembryonic antigen(CEA),cancer antigen(CA)125 and CA19-9 combined tests for liver metastasis is unclear.AIM To evaluate predictive value of combined tests for CEA,CA125,and CA19-9 levels in patients with liver metastases of CRC.METHODS The retrospective study included patients with CRC alone(50 cases)and patients with CRC combined with liver metastases(50 cases)who were hospitalized between January 2021 and January 2023.Serum CEA,CA125 and CA19-9 levels were compared between the two groups,and binary logistic regression was used to analyze the predictive value of the combination of these tumor markers in liver metastasis.In addition,we performed receiver operating characteristic(ROC)curve analysis to assess its diagnostic accuracy.RESULTS The results showed that the serum CEA,CA125 and CA19-9 levels in the CRC with liver metastasis group were significantly higher than those in the CRC alone group.Specifically,the average serum CEA level in the CRC with liver metastasis group was 162.03±810.01 ng/mL,while that in the CRC alone group was 5.71±9.76 ng/mL;the average serum CA125 levels were 43.47±83.52 U/mL respectively.and 13.5±19.68 U/mL;the average serum CA19-9 levels were 184.46±473.13 U/mL and 26.55±43.96 U/mL respectively.In addition,binary logistic regression analysis showed that CA125 was significant in predicting CRC liver metastasis(P<0.05).ROC curve analysis results showed that the areas under the ROC curves of CEA,CA125 and CA19-9 were 0.607,0.692 and 0.586.CONCLUSION These results suggest that combined detection of these tumor markers may help early detection and intervention of CRC liver metastasis,thereby improving patient prognosis.

Significance of carcinoembryonic antigen detection in the early diagnosis of colorectal cancer:A systematic review and metaanalysis

BACKGROUND Colorectal cancer(CRC)is a prevalent malignant tumor involving adenomas that develop into malignant lesions.Carcinoembryonic antigen(CEA)is a non-specific serum biomarker upregulated in CRC.The concentration of CEA is modulated by tumor stage and grade,tumor site in the colon,ploidy status,and patient smoking status.This study aimed to evaluate current evidence regarding the diagnostic power of CEA levels in the early detection of CRC recurrence in adults.AIM To evaluate current evidence regarding the diagnostic power of CEA levels in the early detection of CRC recurrence in adults.METHODS A systematic search was performed using four databases:MEDLINE,Cochrane Trials,EMBASE,and the Web of Science.The inclusion criteria were as follows:Adult patients aged≥18 years who had completed CRC curative treatment and were followed up postoperatively;reporting the number of CRC recurrences as an outcome;and randomized,clinical,cohort,and case-control study designs.Studies that were not published in English and animal studies were excluded.The following data were extracted by three independent reviewers:Study design,index tests,follow-up,patient characteristics,and primary outcomes.All statistical analyses were performed using the RevMan 5.4.1.RESULTS A total of 3232 studies were identified,with 73 remaining following the elimination of duplicates.After screening on predetermined criteria,12 studies were included in the final analysis.At a reference standard of 5 mg/L,CEA detected only approximately half of recurrent CRCs,with a pooled sensitivity of 59%(range,33%–83%)and sensitivity of 89%(range,58%–97%).CONCLUSION CEA is a significant marker for CRC diagnosis.However,it has insufficient sensitivity and specificity to be used as a single biomarker of early CRC recurrence,with an essential proportion of false negatives.

False positive detection of serum cryptococcal antigens due to insufficient sample dilution:A case series

At present,with the development of technology,the detection of cryptococcal antigen(CRAG)plays an increasingly important role in the diagnosis of cryptococcosis.However,the three major CRAG detection technologies,latex agglutination test(LA),lateral flow assay(LFA)and Enzyme-linked Immunosorbent Assay,have certain limitations.Although these techniques do not often lead to false-positive results,once this result occurs in a particular group of patients(such as human immunodeficiency virus patients),it might lead to severe consequences.

Comparison of Two Enzyme Immunoassays and Four Lysate Antigens for the Detection of Antibody in Canine Blastomycosis

Blastomycosis, the systemic fungal disease of humans and animals caused by Blastomyces dermatitidis and the cryptic species Blastomyces gilchristii, is often misdiagnosed as a bacterial or viral pulmonary disease. Therefore, the development of improved immunodiagnostic assays for this disease has been the primary focus of research in our laboratory. The present study was designed to evaluate four Blastomyces yeast-phase lysate antigenic preparations (human, 597, Eagle River, WI;dog, ERC-2, WI;Human, B5927, Mountain Iron, MN;soil, 85, Georgia, ATCC 56920) for their ability to detect antibody in 48 serum specimens from dogs with diagnosed blastomycosis using an indirect ELISA (STD) compared to a biotin-streptavidin ELISA (B-SA). All four lysate antigens were able to detect antibodies in the specimens with mean absorbance values ranging from 0.930 (B5927) to 1.142 (ERC-2) with the STD ELSA and from 1.395 (B5927) to 1.775 (85) with the B-SA ELISA. The results indicated that both ELISA methods could be utilized for antibody detection, but the B-SA ELISA exhibited greater sensitivity than the STD ELISA with all four of the lysates.

Clinical significance of the detection of Rh blood group antigens and irregular antibodies in pregnant women with a second pregnancy

Objective:To investigate the phenotype distribution of five antigens of Rh blood group system and the specificity of Rh blood group irregular antibodies in pregnant women with second child.To analyze the relationship between Rh blood group antibody and hemolytic disease of the newborn(HDN)in second-child pregnant women,and to provide laboratory basis for the diagnosis and treatment of hemolytic disease of the newborn(Rh-HDN).Methods:500 pregnant women with second child were collected as the study group and 500 pregnant women with first pregnancy as the control group(all pregnant women underwent obstetric examination in the integrated obsteric clinic of our hospital from January 2020 to January 2021).To detectethe Rh blood group antigens(D,C,c,E,e)of the two groups of samples,screene the irregular antibodies,identify the specificity of irregular antibodies,determine the titer and record the hemolytic disease of the newborn of pregnant women with positive Rh blood group antibodies.Results:There were 11 Rh phenotypes in the pregnant women with second child in the study group:CCDee(152cases,30.4%),CcDEe(136cases,27.2%)CcDee(84cases,16.8%),ccDEE(30cases,6%),ccDee(31cases,6.2%),CCDEe(14cases,2.8%),ccDEe(9cases,1.8%),cc dee(18cases,3.6%),CCDEE(2cases,0.4%),CcdEe(12cases,2.4%),Ccdee(6cases,1.2%),CCd ee(6cases,1.2%).A total of 42 cases(8.4%)in the pregnant women with second child were negative for RhD.There were 10 Rh phenotypes in the pregnant women with first pregnancy in the control group:CCDee(144cases,28.8%),CcDEe(138cases,27.6%),CcDee(90cases,18%),ccDEE(42cases,8.4%),ccDee(28cases,5.6%),CCDEe(10cases,2%),ccDEe(8cases,1.6%),cc dee(19cases,3.8%),CCDEE(1cases,0.2%),CcdEe(11cases,2.2%),Ccdee(9cases,1.8%).A total of 39 cases(7.8%)in the pregnant women with first pregnancy were negative for RhD.In the pregnant women with second child in the study group,the positive rate of irregular antibody screening was 4.0%(20/500),and the specificity of Rh blood group antibodies was found as follows:anti-E 1.8%(9/500),anti-D 1.4%(7/500),anti-C 0.4%(2/500)and anti-Ec 0.4%(2/500).The positive rate of irregular antibody screening in the pregnant women with first pregnancy in the control group was 0,and the difference between the two groups was statistically significant(P<0.05).Rh-HDN was found in 10 newborns(2%)of the 20 women with positive irregular antibodies in the pregnant women with second child,and the antibody titer during pregnancy was more than 32.No Rh-HDN occurred in newborns in the pregnant women with first pregnancy,and the difference between the two groups was statistically significant(P<0.05).Conclusion:Pregnancy stimulation can increase the probability of irregular antibodies in pregnant women,and irregular antibodies in Rh blood group can easily cause Rh-HDN,so attention should be paid to routine detection of five antigens of Rh blood group and irregular antibody screening during prenatal examination.It is helpful for the early detection of Rh-blood irregular antibodies and the assessment of fetal or neonatal risk of Rh-HDN.

GENE ENGINEERING EB VIRUS MEMBRANE ANTIGEN IN DETECTION OF MA-IgA ANTIBODY(COMPARISON WITH VCA-IgA AND EA-IgA ANTIBODIES)

With gene engineering EB virus membrane antigen as the diagnostic antigen, indirect immunofluo-rescence (IF) assay was used to detect IgA antibody against EB virus membrane antigen (MA-IgA) in sera from 202 nasopharyngeal carcinoma (NPC) patients and 315 controls (normal and patients with other tumors). MA-IgA antibody was positive in 96.8% of the pretreatment NPC patients with a GMT of 1:36.3. MA-IgA detection by this method was more sensitive than EA-IgA detection by IE. In contrast, patients with tumors other than NPC were negative for MA-IgA antibody. 9.1% of VCA-IgA positive persons were MA-IgA positive with a GMT of less than 1:5. No MA-IgA positive was found in VCA-IgA negatives. The results indicated that this method was relatively specific. In the treatment group, the positive rate and GMT of MA-IgA antibody declined with increase in survival time and the decline was faster than VCA-IgA. When recurrence or distant metastasis developed, similar to VCA-IgA and EA-IgA antibodies, the positive rate and GMT of MA-IgA antibody increased to its pretreatment level. Therefore, MA-IgA detection might be valuable in the early diagnosis and monitor of NPC.

Fast Electrical Detection of Carcinoembryonic Antigen Based on AlGaN/GaN High Electron Mobility Transistor Aptasensor

As one of the most important tumor-associated antigens of colorectal adenocarcinoma, the carcinoembryonic antigen （CEA） threatens human health seriously ali over the globe. Fast electrical and highly sensitive detection of the CEA with A1GaN/GaN high electron mobility transistor is demonstrated experimentally. To achieve a low detection limit, the Au-gated sensing area of the sensor is functionalized with a CEA aptamer instead of the corresponding antibody. The proposed aptasensor has successfully detected different concentrations （ranging from 50picogram/milliliter （pg/ml） to 50 nanogram/milliliter （ng/ml）） of CEA and achieved a detection limit as low as 50pg/ml at Vas = 0.5 V. The drain-source current shows a c/ear increase of 11.5μA under this bias.

Critical Considerations in the Immunochemical Detection and Quantitation of Antigenic Biomarkers

The formation of covalent adducts as a result of the interaction of metabolically activated chemicals with host macromolecules is a common critical event in mutagenic, carcinogenic, and immunologic phenomena. Because of their antigenicity and their immunogenicity, covalent adducts may be detected using sensitive immunochemical techniques. The immunochemical approaches to biomonitoring and molecular dosimetry of DNA damage are particularly attractive because they allow sensitive quantitation of specific DNA adducts present in small samples and do not rely on the use of radiolabeled adducts. Two examples of biomarker immunoassay development are presented: an avidin/biotin-amplified ELISA for the major DNA adduct of the human bladder carcinogen 4-aminobiphenyl (ABP), and a particle concentration fluorescent immunoassay (PCFIA) for the major protein adduct associated with toxicity by the prototype hepatotoxin acetaminophen. The examples illustrate critical steps in the development of biomarker immunoassays which include selection of the relevant adduct, preparation of an appropriate immunogen, immunization, characterization of antisera, and development of application-specific sample processing techniques for biomarker quantitation. Immunochemical procedures may be combined with other analytical techniques to form hybrid systems which take advantage of both the antigenicity and the physical or chemical properties of a biomarker to achieve greater specificity and/or sensitivity. The future usefulness of these new tools of molecular epidemiology will depend on a compound-by-compound validation of methods and critical evaluation of the biologic importance of the particular antigenic biomarker as an indicator of exposure and as an indicator of risk.

Application of an indirect immunofluorescent staining method for detection of Salmonella enteritidis in paraffin slices and antigen location in infected duck tissues

AIM： To detect Salmonella enteritidis （S. enteritidis） in paraffin slices and antigen location in infected duck tissues. METHODS： The rabbits were immunized with purified bacillus to obtain S. enteritidis-specific antibody, which were then extracted by the caprylic-ammonium sulphate method, purified through High-Q columns. An indirect immuno-fluorescent staining method （IFA） was established to detect the S. enteritidis antigen in paraffin slices. Detected S. enteritidis in each organ tissue of ducklings experimentally infected with S. enteritidis. RESULTS： The gland of Garder, heart, kidney, spleen, liver, brain, ileum, jejunum, bursa of Fabricius from S. enteritidis experimentally infected ducklings were positive or strongly positive, and the S. enteritidis antigen mainly distributed in the infected cell cytoplasm.CONCLUSION： IFA is an intuitionist, sensitive and specific method in detecting S. enteritidis antigen in paraffin wax slices, and it is a good method in diagnosis and antigen location of S. enteritidis. We also conclude that the gland of Garder, heart, kidney, spleen, liver, ileum, jejunum are target organs in S. enteritidis infections of duck, and S. enteritidis is an intracellular parasitic bacterium.

Antigen Detection in Canine Blastomycosis: Comparison of Different Antibody-Antigen Combinations in Two Competitive ELISAs

This present study was designed to evaluate four different Blastomyces dermatitidis antibody-antigen combinations (B5896 and T-58 antibodies and B5896 and WI-R antigens) for the detection of antigen in 36 urine specimens from dogs with blastomycosis using a standard indirect ELISA (STD) and a biotin-streptavidin ELISA (B-SA). The antigen detection sensitivity values ranged from 81% (B-SA: T-58 Ab + WI-R Ag) to 100% (STD and B-SA: B5896 Ab + WI-R Ag;B5896 Ab + B5896 Ag) with the antibody-antigen combinations in the two assays. Optimal detection was evidenced when the B5896 Ab was allowed to react with the urine specimens for 30 min at 37?C and then placed in the B-SA ELISA plates containing the B5896 Ag. The greatest absorbance value obtained with this antibody-antigen com-bination was 0.903 (range of 0.596 - 0.903) as compared to the control value of 1.246. The difference between the control absorbance and the test absorbance values was 0.343 which was considerably greater than the control-test values with the other combinations. This study thus showed that the results obtained in antigen detection assays are dependent upon the antibody used to react with the urine specimens as well as the antigen used in the enzyme immunoassay.

Comparative antibody study for antigen detection in urine specimens for diagnosis of blastomycosis using a competitive enzyme-linked immunosorbent assay

Diagnosis of blastomycosis is often done using a combination of clinical signs and cytologic or histopathologic identification of the organism, Blastomyces dermatitidis, from infected tissues. However, these methods are time consuming, invasive, and still lead to misdiagnosis. A competitive enzyme-linked immunosorbent assay (ELISA) can be used for detection of B. dermatitidis antigens, which are present in urine specimens of infected patients. The current study evaluates the use of various antibodies for detection of antigen in dog urine specimens, to provide a better diagnosis of blastomycosis in the future. Our results show that different antibodies against B. dermatitidis produce various sensitivities for antigen detection. The most realistic antibodies for immunodiagnostic tests would be antibodies that can be obtained in larger quantities, i.e. vaccination using a yeast lysate in a laboratory setting. We found that these antibodies produce a comparable and reliable result to that of antibodies obtained from an infected patient.

Value of carcinoembryonic antigen and cytokeratins for the detection of recurrent disease following curative resection of colorectal cancer

AIM： To evaluate the efficacy of postoperative serial assay of carcinoembryonic antigen （CEA） and cytokeratins for the detection of recurrent disease in patients with colorectal adenocarcinoma after radical surgery. METHODS： Between 1993 and 2000, 120 patients with colorectal adenocarcinoma underwent radical surgery in the Department of Surgical Gastroenterology, Federal University of Sao Paulo-Escola Paulista de Medicina, Sao Paulo, Brazil. Periodic postoperative evaluation was performed by assaying markers in peripheral serum, colonoscopy and imaging examination. Presence of CEA was detected using the Delfia^R method with 5 μg/L threshold, and cytokeratins using the LIA-mat TPA-M Prolifigen^R method with 72 U/L threshold. RESULTS： In the first postoperative year, patients without recurrent disease had normal levels of CEA （1.5 ＋ 0.9μg/L） and monoclonal tissue polypeptide antigen-M （TPA-M, 64.4 ± 47.8 U/L）, while patients with recurrences had high levels of CEA （6.9± 9.8 ;μg/L, P 〈 0.01） and TPA-M （192.2 ±328.8 U/L, P 〈 0.05）. During the second postoperative year, patients without tumor recurrence had normal levels of CEA （2.0 ± 1.8μg/L） and TPA-M （50.8±38.4 U/L）, while patients with recurrence had high levels of CEA （66.3 ±130.8 μg/L, P 〈 0.01） and TPA-M （442.7 ± 652.8 U/L, P 〈 0.05）. The mean follow-up time was 22.3 mo. There was recurrence in 23 cases. Five reoperations were performed without achieving radical excision. Rises in tumor marker levels preceded identification of recurrences： CEA in seven （30%） and TPA-M in eleven individuals （48%）. CONCLUSION： Intensive follow-up by serial assay of CEA and cytokeratins allows early detection of colorectal neoplasm recurrence.

Role of serum carcinoembryonic antigen in the detection of colorectal cancer before and after surgical resection

AIM:To determine whether serum levels of carcinoembryonic antigen(CEA) correlate with the presence of primary colorectal cancer(CRC),and/or recurrent CRC following radical resection.METHODS:A total of 413 patients with CRC underwent radical surgery between January 1998 and December 2002 in our department and were enrolled in this study.The median follow-up period was 69 mo(range,3-118 mo),and CRC recurrence was experienced by 90/413(21.8%) patients.Serum levels of CEA were assayed preoperatively,and using a cutoff value of 5 ng/mL,patients were divided into two groups,those with normal serum CEA levels(e.g.,≤ 5 ng/mL) and those with elevated CEA levels(> 5 ng/mL).RESULTS:The overall sensitivity of CEA for the detection of primary CRC was 37.0%.The sensitivity of CEA according to stage,was 21.4%,38.9%,and 41.7% for stagesⅠ-Ⅲ,respectively.Moreover,for stageⅡandstageⅢcases,the 5-year disease-free survival rates were reduced for patients with elevated preoperative serum CEA levels(P < 0.05).The overall sensitivity of CEA for detecting recurrent CRC was 54.4%,and sensitivity rates of 36.6%,66.7%,and 75.0% were associated with cases of local recurrence,single metastasis,and multiple metastases,respectively.In patients with normal serum levels of CEA preoperatively,the sensitivity of CEA for detecting recurrence was reduced compared with patients having a history of elevated CEA prior to radical resection(32.6% vs 77.3%,respectively,P < 0.05).CONCLUSION:CRC patients with normal serum CEA levels prior to resection maintained these levels during CRC recurrence,especially in cases of local recurrence vs cases of metastasis.

Analysis of Commercial Assays for the Detection of SARS-CoV-2 Antibodies or Antigens

Background: COVID-19 produced by SARS-CoV-2 infection has spread worldwide. There is a growing need for immunological assays to detect viral specific antibodies or viral antigen. Current standard of diagnosis is reverse-transcriptase polymerase chain reaction (RT-PCR) in nasopharyngeal swabs. However serological tests can be used to determine previous exposure to the virus and complement the diagnosis. IgM and IgG SARS-CoV-2 specific antibodies can be detected as early as one week after infection and assays can be useful to test large groups of individuals. This work revised the available information concerning assays that detect antibodies and antigens for SARS-CoV-2. Methods: Three sources of information were used: technical data sheets (TDS) web pages of the company’s products and published articles in Pubmed with reference to the use of diagnostic kits. All the information was revised until April 5th 2020. Results: There were 226 tests coming from 20 countries mainly from China. TDS were found only in 50 (22.1%). Most assays detect specific antibodies (n 180) based on immunochromatography methods (n 110) and use blood-derived samples (n 105). Assays for antibodies detection measured mainly IgM/IgG (n 112) and the most common procedure time was <20 min (n 83). Internal control referred as sensitivity and specificity was found only in 18.6% (n 42) of the assays. The majority of the tests are currently for in vitro diagnosis (IVD). A total 165 articles were found on PubMed 15 were included and only 4 used the commercial kits reviewed. Conclusions: Due to the urgency of producing diagnostic tests for SARS-CoV-2 there is a broad offer of kits. Many tests need additional information for their application. The data collected may be useful in the selection of assays but more and higher quality information is needed.

Evaluation of an Innovative Diagnostic Method for Detection of Antibodies and Antigens

Reports manifest a continuing need for the development of rapid and on-site (point of care) assays. Current diagnostic methods commonly used for detection of antibodies and antigens have significant limitations. Scientists at Micro Detect, Inc. have developed an innovative diagnostic device (method) that can be utilized broadly for antibody/antigen interactions including diagnostic assays in the medical, veterinary and food industries. The developed device can be utilized for the detection of antibodies against a single antigen or vice versa. It can also be tailored for specific panels that detect antigens or antibodies for diverse infectious agents, proteins, hormones, tumor markers, autoimmune markers, and allergens. Additionally, it can also be used for detection of toxins, antitoxins, nucleic acids, enzymes, drugs, etc. in both humans and animals. Specimens used in different formats of the device can be tears, saliva, whole blood, serum, plasma, urine, stool, and other bodily discharges. The good intra and inter precisions and acceptable linearity of the device support reliable use of the device. The CV of the device is 1.9% - 2.2%. Likewise, the performance of the device using 92 confirmed negative and positive specimens via a typical assay showed 100% sensitivity, 80% specificity, 96.8% efficacy, 80% positive predictive value, and 100% negative predictive value. The results of our feasibility study suggest reliable utility of a device for rapid, easy-to-use, inexpensive, and on-site (point of care) diagnostic assays. This presents a potential breakthrough in diagnostic methodologies that can be integrated into modern medicine and food industries.

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.

Comparison of Antibody Detection with Yeast Lysate Antigens Prepared from Blastomyces dermatitidis Dog Isolates from Wisconsin and Tennessee

Blastomyces dermatitidis, the causative agent of blastomycosis, a potentially lethal dimorphic fungal disease of humans and animals has been difficult to diagnose in the clinical laboratory. We are attempting to develop and improve immunodiagnostic assays by producing novel yeast lysate reagents for the detection of antibodies in blastomycosis. The objective of this study was to use lysate antigens prepared from four B. dermatitidis antigens isolated from dogs infected with blastomycosis from two different endemic areas (Wisconsin and Tennessee) testing for the detection of antibodies in serum specimens from immunized rabbits and infected dogs using the indirect ELISA. In the dog sera, absorbance values ranged from 0.774 to 1.350, while the rabbit sera values ranged from 0.533 to 1.191. Antigen T-58 appeared to lack any geographical specificity in antibody detection, which could prove useful in future immunodiagnostic detection of blastomycosis infections.

Comparison of antibody detection with <i>Blastomycesdermatitidis</i>yeast lysate antigens in serum specimens from immunized rabbits and infected dogs

This present study was designed to evaluate B. dermatitidis antigens, prepared from two isolates (B5896, 597), when the yeast cells were allowed to lyse in distilled water for one day or seven days. The indirect enzyme-linked immunosorbent assay (ELISA) was used to determine the ability of the lysate reagents to detect antibodies in 30 rabbit and 30 dog serum specimens. Mean absorbance values with B5896 lysate antigen ranged from 1.637 (day 1) to 1.461 (day 7) and absorbance values with 597 antigen ranged from 1.579(day 1) to 1.396 (day 7) with the serum specimens from immunized rabbits. Serum specimens from infected dogs yielded absorbance values ranging from 1.672 (day 1) to 1.763 (day 7) with the B5896 and values ranging from 1.909 (day 1) to 1.224 (day 7) with the 597. Optimal reactivity was obtained with the day 1 lysate using both lysate antigens against the rabbit sera and with the 597 antigen against the dog sera. Slightly greater reactivity was evidenced with the day 7 B5896 antigen when the dog sera was tested. Comparative studies are continuing in order to produce an optimal anti-genic preparation for antibody detection in blastomycosis.