Contribution of Anti-p63 Antibodies in the Interpretation of Benign Label Prostatic Biopsies

Introduction: Prostate cancer is the second most common cancer in men. The diagnosis is most often based on the prostate biopsies’ analysis and on histological criteria recognizable on standard coloring. In some cases, the use of immunohistochemistry is important. Objectives: This paper aims to specify the p63 phenotypic profile of lesions diagnosed benign, with minimal suspect foci, difficult to interpret, HGPIN (high grade intraepithelial neoplasia) and LGPIN (low-grade prostatic intraepithelial neoplasia) and evaluate the manual technique of p63 immunohistochemistry. Patients and Method: This was a retrospective, descriptive study of prostate biopsies recorded in the PAC service of the HALD from January 1st, 2018 to December 31st, 2018. It was completed by a manual immunohistochemical study of the blocks enrolled from November 19th to December 4th, 2020 in the PAC department of the HPD. The studied parameters were: registry number, age, clinical stage, prostate volume, PSA level, microscopic appearance and p63 immunohistochemical profile. Results: Our study included 60 prostate biopsies. The ages of our patients varied from 45 to 77 years, with an average of 64.2 years and a standard deviation of 6.2. The majority of patients were at clinical stage cT2b (33%) with a prostate volume varying between 33.15 and 169.4 cc. The minimum value of PSA in our series is 5 ng/ml, the maximum being 100 ng/ml with an average level of 24.1 ng/ml and a standard deviation of 21.2. Our series included 50 adenomyomatous hyperplasias, 7 adenomyomatous hyperplasias associated with chronic prostatitis, 2 HGPIN and 1 LGPIN. After re-reading we found 5 discordant cases, which corresponded to minimal suspect foci (kappa = 0.5098). The p63 marking was informative in 53 cases, i.e. 88%, and non-informative in 7 cases, i.e. 12%. Among the uninformative markings, 2 were due to lack of tissue adhesion to the slides. Among the informative markings, 11 were negative. p63 immunohistochemistry was useful in all suspected foci and detected 6 other minimal foci of adenocarcinoma. Conclusion: The immunostaining with the anti-p63 antibody in the prostate cancer diagnosis is of considerable benefit. It made it possible to correct 11.3% of benign diagnosis in minimal malignant focus in our context. Despite the difficulties associated with the manual technique, it is possible to have an informative rate, similar to the automatic technique.

CLINICAL EVALUATION OF MONOCLONAL ANTIBODIES AGAINST ALDOLASE-A IN THE DIAGNOSIS OF HEPATO-CELLULAR CARCINOMA

Serum ALD-A of 57 patients with HCC and 43 of other diseases were measured by ALD-A-McAb linked with 425I-staphylococcus A protein. The results showed that the ALD-A in patient with HCC was significantly elevated as compared with controls and that in patients with cholangiocarcinoma, gastrointestinal cancer without hepatic metastasis, cirrhosis. CAH and benign GI diseases. There was no statistical difference between ALD-A in patients with HCC and that in cases of cirrhosis with liver failure and that in cases of metastatic liver carcinoma. It was noted that diagnostic sensitivity of ALD-A in AFP (+) HCC was 73.9% and that in AFP (-) HCC was 81.8% 1-6 patients with HCC were treated by hepatic arterial embolization combined with chemotherapy. ALD-A in patients after the treatment decreased significantly than that before treatment, furthermore, advantages of the method are discussed.

DETECTION OF CANCER-ASSOCIATED ANTIGEN IN FECES USING MONOCLONAL ANTIBODIES IN THE DIAGNOSIS OF COLON CARCINOMA

Monoclonal antibodies against colon and pancreatic cancer, CL-2, CL-3, PS-9, PS-10, were used to detect the associated antigens in feces of patients with gastrointestinal carcinoma and non-cancer diseases. Binding inhibition test by SABC-ELISA method were performed for the measurement of the antigen level. Results showed that the associated antigen detected in feces of patients with colon cancer were significantly higher than that of non-cancer disease or normal subjects. The positive rates were 61.1% as detected with CL-2; 53.4% with CL-3; 55.0%, PS-9; and 53.3% PS-10 in cancer patients while that in normal subjects were 7%; 9%; 8%; and 8% respectively. When 'cocktail' of CL-2, PS-9 and PS-10 were used, the positive rates were 92.5% in colon cancer and 14% in normal subjects. In seven out of the sixty patients with colon cancer studied who were graded as Dukes A, the results were all positive. The results seem superior to the serologic detection and may provide a promising new approach in the early diagnosis of colon cancer.

Translational research case studies:Development of antibodies for cancer diagnosis and therapy

Antibodies are primary tools in several areas of biomedical sciences, including basic research, diagnostics, and molecular therapeutics. Antibodies are widely used in diagnostic applications for clinical medicine. Analysis of cells and tissues in pathology laboratories includes the use of antibodies on tissue sections. Further, antibodies are making rapid inroads into medical therapeutics, driven by technological evolution from chimeric and humanized to fully human antibodies. The therapeutic antibody market has the potential to reach $30 billion by 2010. Our lab has developed a monoclonal antibody, named Met4 that was raised against the extracellular domain of Met specifically with the goal of measuring Met in FFPE tissues. The Met receptor kinase is expressed on the cell surface of a significant number and variety of human primary solid tumors and in their metastases. The characterization of the Met4 antibody suggests it should possess adequate performance for quantification of Met expression in clinical specimens. We have also generated a fully human Fab fragment against EGFR; conjugated it to taxol as an immuno-chemotherapy agent; and investigated its in vitro antitumor efficacy on EGFR positive A431 epidermoid carcinoma cells.

Endomysial antibodies predict celiac disease irrespective of the titers or clinical presentation

AIM:To investigate the association between serum antibody levels and a subsequent celiac disease diag-nosis in a large series of children and adults. METHODS:Besides subjects with classical gastrointestinal presentation of celiac disease,the study cohort included a substantial number of individuals with extraintestinal symptoms and those found by screening in at-risk groups.Altogether 405 patients underwent clinical,serological and histological evaluations.After collection of data,the antibody values were further graded as low[endomysial(EmA)1:5-200,transglutaminase 2 antibodies(TG2-ab)5.0-30.0 U/L]and high (EmA 1:≥500,TG2-ab≥30.0 U/L),and the serological results were compared with the small intestinal mucosal histology and clinical presentation. RESULTS:In total,79%of the subjects with low and 94%of those with high serum EmA titers showed small-bowel mucosal villous atrophy.Furthermore, 96%of the 47 EmA positive subjects who had normal mucosal villi and remained on follow-up either subsequently developed mucosal atrophy while on a glutencontaining diet,or responded positively to a glutenfree diet. CONCLUSION:Irrespective of the initial serum titers or clinical presentation,EmA positivity as such is a very strong predictor of a subsequent celiac disease diagnosis.

Diagnostic role of antibodies to glutamic acid decarboxylase in latent autoimmune diabetes mellitus in adults

Objective To investigate the diagnostic role of antibodies to glutamic acid decarboxylase (GAD 65 Ab) in latent autoimmune diabetes of adults (LADA) and the frequency of GAD Ab in Chinese patients initially diagnosed as non insulin dependent diabetes mellitus (NIDDM) Methods Forty five control subjects and 195 consecutive inpatients initially classified as NIDDM with ≥35 years of age at onset and nonketotic history for >6 months after diagnosis, were recruited In vitro transcripted and translated recombinant human 35 S GAD 65 was used in radioligand assay of GAD Ab Results The overall prevalence of GAD 65 Ab was 14 8% (29/195) in NIDDM patients and 2 2% (1/45) in control subjects, respectively Of the 29 GAD 65 Ab positive patients, 17 (58 6%) were insulin deficient while 12 (41 4%) were non insulin deficient The prevalence of GAD 65 Ab in NIDDM group with age of <40 years at diabetes onset, ketotic history, body mass index (BMI) <21 kg/m 2, were significantly higher than that of corresponding control diabetic subgroups (2 5, 4 1 and 3 2 times, respectively) The sex, duration, symptoms of polyphagia, polydipsia, polyuria and weight loss at onset of the disease were not related to the prevalence of GAD 65 Ab positivity Conclusions In China, patients initially diagnosed as NIDDM may in many cases suffer from LADA Testing by GAD 65 Ab may be of assistance to identifying LADA at the earliest stage of disease

Novel P22-monoclonal antibody based blocking ELISA for the detection of African swine fever virus antibodies in serum

African swine fever(ASF)is a highly infectious,transboundary viral disease of domestic and wild pigs,and is currently the most serious threat to world swine production,resulting in significant economic loss.In the absence of vaccines and treatments,the control of the disease entirely depends on accurate and early diagnosis accompanied by the culling of infected pigs.Thus,a highly specific and sensitive diagnostic assay is required during an outbreak and surveillance of the disease.In this study,a highly sensitive,specific,rapid and repeatable P22-monoclonal antibody-based blocking enzyme-linked immunosorbent assay(bELISA)assay was developed for the detection of antibodies against genotype I and II African swine fever viruses(ASFVs).A total of 806 pig serum samples were tested to evaluate the performance of the diagnostic assay.To determine the PI(percent Inhibition)cut-off value,receiver-operating characteristic(ROC)analysis was applied.According to the ROC analysis of the data,98.10%specificity and 100%sensitivity were recorded when the threshold cut-off value of PI was established at 47%.In addition,the assay was able to detect ASFV antibodies as early as 9 days post-infection when serum samples from experimentally infected pigs were used.Taking all together,the results of the present study indicated that the P22-mAb based bELISA assay can be used for rapid and accurate detection of antibodies against ASFV,which could play a valuable role in the containment and prevention of ASFV as an alternative to other serological diagnostic methods.Also,this study will assist researchers to further investigate the immunogenic importance of P22 protein in ASFV infection.

Autoantibodies in psoriatic arthritis:are they of pathogenic relevance?

A physician in clinical practice does not usually order autoantibody testing to aid subsequent diagnosis or for monitoring disease activity in patients with psoriasis or psoriatic arthritis(PsA),although a variety of autoantibodies are present in these patients.Our understanding of autoantibodies in psoriasis and PsA is limited.Early investigations of autoantibodies in psoriasis were focused on the known autoantibodies in rheumatic diseases.For instance,anti-nuclear antibodies(ANAs)are often found in patients with psoriasis or PsA,but anti-double-stranded DNA or anti-extractable nuclear antigens are rarely identified.Therefore,ANAs have not been considered valuable in diagnosing PsA or predicting prognosis to manage PsA.Moreover,the roles of ANAs in the pathogenesis of PsA remain unknown.[1,2]Autoantibodies associated with rheumatoid arthritis(RA)have been investigated in PsA for their presence and association with the disease.For instance,antibodies against citrullinated proteins(ACPAs),which are highly specific to RA,are found in 5.0%to 17.5%of PsA patients.In several studies,a more erosive disease has been observed in PsA patients with ACPAs than in ACPA-negative PsA patients.[3,4]These findings imply that ACPAs in patients with PsA may be capable of inducing bone loss,which has been observed in RA patients with antibodies against citrullinated vimentin.

Comparison of Multiplex Fluorescent PCR with Serum Type-specific Antibody Detection in Diagnosis of Genital Herpes

Objectives: To compare multiplex fluorescent PCRwith serum type-specific antibody detection in thediagnosis of herpes simplex virus (HSV) infection andto evaluate its significance in the diagnosis of genitalherpes.Methods: We detected HSV infection in 121 speci-mens collected from patients with genital herpesusing both multiplex fluorescent PCR and serum type-specific antibody detection. HSV viral isolation wasused as the standard control.Results: When compared with the viral isolation, thesensitivity and specificity for multiplex fluorescentPCR were 100% and 88.89%, respectively afterdiscrepant analysis. The sensitivity and specificity fortype-specific antibody detection was 77.68 % and77.78 %, respectively. However, the type-specificantibody detected HSV in two asymptomatic patientswhile the multiplex fluorescent PCR couldn’t detectany HSV DNA from those specimens.Conclusions: Multiplex fluorescent PCR is a verysensitive and specific method for detection and typingof HSV in the lesion of genital herpes, it failed todetect HSV DNA from the asymptomatic patients.Serum type-specific antibody detection was a lesssensitive and specific test but could detect the specificantibody from some asymptomatic patients. Thecombination of these two techniques would allow rapid,sensitive and accurate detection and typing of HSVand help clinical diagnosis and epidemiologic survey-ing of genital herpes.

STUDIES ON SPECIFIC SEROLOGICAL ANTIGENS IN METACERCARIAE AND JUVENILES OF Paragoniums westermani AND ITS MONOCLONAL ANTIBODIES

The polyclonal antibodies to juveniles of Paragoniums westermani (PwJ-PcAbs) from sera of Wistar rats infected with Paragoniums westerrnani (P. w.)were purified by Sephadex G 200 chromatography. Next the shared serological antigens of P. w. metacercaria and juveniles (PwMJ-SAg) from the crude antigens of the metacercariae (M-NS-Ag) were purified with immuno-affinity chromatography on cyanogen bromide-activated cross-linked Sepharose 4B beads coupled withPwJ-PcAbs. PwMJ-SAg, agroup of glycoprotein molecules shown by the staining test, were specific serological antigens of P. w. metacercariae and juveniles, identified by the immunoabsorb test and immunoelectrophoresis. By SDSPAGE, PwMJ-SAg were fractionated to seven bands, including major bands A(27.SK) and Bi(19.5 K), the two major serological antigen molecules. 20 sera samples from the patients with the nonpulmonary type of P. w. paragonimiasis were detected using PwMJ-SAg and M-NS-Ag by Dot-ELISA, and the difference of sensitivity between two antigens was highly statistically significant (P<0.001). BALB/c mice, in the early stage of infection with P. w. metacercaria, were immunized with PwMJ-SAg. The spleen cells of the mice were isolated and fused with SP_2/o, a murine myeloma cell line. After three subclonal cultures, eight cell lines secreting monoclonal antibodies (McAbs) to PwMJ-SAg were prepared from 384 wells of hybridoma cells. All McAbs were IgG_1 subclass.

Electrochemical Immunosensors for the Diagnosis of Celiac Disease

Celiac disease is a permanent intolerance to gluten proteins of wheat, rye, barley, and oats in genetically susceptible individuals. The clinical picture is characterized by inflammation and damage of the small intestinal mucosa and malabsorption of essential nutrients. Therapeutically, a lifelong strict gluten-free diet is necessary. The diagnosis of celiac disease is complex and includes symptomatology, serology, small intestinal histology, and genetic status. Serological testing plays a central role within the diagnostic procedure and is based on the measurement of disease-specific antibodies against gluten proteins (antigen) and tissue transglutaminase (autoantigen). Immunofluorescence detection and enzyme-linked immunosorbent assays are currently most often applied for antibody testing. However, these tests are expensive and time-consuming. Therefore, simple and rapid alternative methods have been developed during the last years, and electro-chemical immunosensors seem to be the most promising analytical tools. The architecture of these sensors may comprise the following elements: working and reference electrodes, covalent or noncovalent binding of the antigen to the surface of the working electrode by means of a functional monolayer, and blocking of unreacted binding sites. The analytical procedure is initiated by adding the analyte (serum antibodies) and an analyte-specific second antibody, which is usually labeled with an enzyme. The special reaction of the enzyme with an appropriate substrate results in a product that initiates a current that can be measured by different electrical methods. A number of different electrochemical immunosensors variable in different electrodes, binding systems, secondary antibodies, and current measurements have been developed. Most of them have been tested with real human serum samples of celiac patients and healthy individuals, and some of them reached disease sensitivity and specificity comparable with traditional analytical systems. Thus, electrochemical immunosensors can be promising alternatives to existing diagnostic tests in the future. They are simple, reliable, robust, user-friendly, and cost-effective tools with short operation times.

Some recent works on diagnosis and treatment of gastric cancer

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Diagnosis of Crohn's disease in India where tuberculosis is widely prevalent

AIM:To define the parameters that positively predict diagnosis of Crohn's disease (CD) and differentiate it from gastrointestinal tuberculosis (GITB). METHODS:This prospective study over 3 years was carried out in the consecutive Indian patients with definite diagnosis of CD and equal numbers of patients with definite diagnosis of GITB. Demographic, clinical, laboratory, morphological and histological features were noted in all the patients. Serological tests such as p-ANCA, c-ANCA, IgA ASCA and IgG ASCA, were performed. Endoscopic biopsy and/or surgical tissue specimens were subjected to smear and culture for acid-fast bacilli (AFB) and tissue polymerase chain reaction for tuberculosis (TB PCR). Diagnosis of CD and GITB was based on the standard criteria. Data were analyzed using univariate Chi-square test and multiple logistic regression (MLR). RESULTS:The study is comprised of 26 patients with CD (age 36.6 ± 8.6 year, male:female, 16:10) and 26 patients with GITB (age 37.2 ± 9.6 year, male:female, 15:11). The following clinical variables between the two groups (CD vs TB) were significant in univariate analysis:duration of symptoms (58.1 ± 9.8 vs 7.2 ± 3.4 mo), diarrhoea (69.2% vs 34.6%), bleeding per rectum (30.7% vs 3.8%), fever (23.1% vs 69.2%), ascites (7.7% vs 34.6%) and extra-intestinal manifestations of inflammatory bowel disease (61.5% vs 23.1%). Of these, all except ascites and extra-colonic manifestations were found statistically significant by MLR. Accuracy of predicting CD was 84.62% based on the fever, bleeding P/R, diarrhoea and duration of symptoms while it was 63.4% when histology was reported as inflammatory bowel disease and 42.3% when there was recurrence of disease after surgery. Accuracy of predicting GITB was 73.1% when there was co-existing pulmonary lesions and/or abdominal lymphadenopathy;75% when tuberculosis was reported in histology;63.4% when granuloma was found in histology;82.6% when TB PCR was positive;and 61.5% when smear and/ or culture was positive for AFB. Serological test was not useful in differentiation of CD from GITB. Positivity rates for CD and GITB were:p-ANCA-3.8% and 3.8%, c-ANCA-3.8% and 0%, IgA ASCA-38.4% and 23.1%, and IgG ASCA-38.4% and 42.3%, respectively. CONCLUSION:Simple clinical parameters like fever, bleeding P/R, diarrhoea and duration of symptoms have the highest accuracy in differentiating CD from GITB.

A comparative laboratory diagnosis of malaria:microscopy versus rapid diagnostic test kits

Objective:To compare the two methods of rapid diagnostic tests(RDTs)and microscopy in the diagnosis of malaria.Methods:RDTs and microscopy were carried out to diagnose malaria. Percentage malaria parasitaemia was calculated on thin films and all non-acute cases of plasmodiasis with less than 0.001%malaria parasitaemia were regarded as negative.Results were simply presented as percentage positive of the total number of patients under study.The results of RDTs were compared to those of microscopy while those of RDTs based on antigen were compared to those of RDTs based on antibody.Patients' follow-up was made for all cases.Results: All the 200 patients under present study tested positive to RDTs based on malaria antibodies(serum)method(100%).128 out of 200 tested positive to RDTs based on malaria antigen(whole blood)method(64%),while 118 out of 200 patients under present study tested positive to visual microscopy of Lieshman and diluted Giemsa(59%).All patients that tested positive to microscopy also tested positive to RDTs based on antigen.All patients on the second day of follow-up were non-febrile and had antimalaria drugs.Conclusions;We conclude based on the present study that the RDTs based on malaria antigen(whole blood)method is as specific as the traditional microscopy and even appears more sensitive than microscopy.The RDTs based on antibody(serum)method is unspecific thus it should not be encouraged.It is most likely that Africa being an endemic region,formation of certain levels of malaria antibody may not be uncommon.The present study also supports the opinion that a good number of febrile cases is not due to malaria. We support WHO's report on cost effectiveness of RDTs but,recommend that only the antigen based method should possibly,be adopted in Africa and other malaria endemic regions of the world.

Evaluation of a Direct Rapid Immunohistochemical Test (dRIT) for Rapid Diagnosis of Rabies in Animals and Humans

Presently the gold standard diagnostic technique for rabies is the direct immunofluorescence assay （dFA） which is very expensive and requires a high level of expertise. There is a need for more economical and user friendly tests, particularly for use in developing countries. We have established one such test called the direct rapid immunohistochemical test （dRIT） for diagnosis of rabies using brain tissue. The test is based on capture of rabies nucleoprotein （N） antigen in brain smears using a cocktail of biotinylated monoclonal antibodies specific for the N protein and color development by streptavidin peroxidase-amino ethyl carbazole and counter staining with haematoxollin. The test was done in parallel with standard FAT dFA using 400 brain samples from different animals and humans. The rabies virus N protein appears under fight microscope as reddish brown particles against a light blue background. There was 100 % correlation between the results obtained by the two tests. Also, interpretation of results by dRIT was easier and only required a light microscope. To conclude, this newly developed dRIT technique promises to be a simple, cost effective diagnostic tool for rabies and will have applicability in field conditions prevalent in developing countries.

Monoclonal antibody and its use in the diagnosis of livestock diseases

Since the discovery of hybridoma cells by Kohler and Milstein, the uses of monoclonal antibody (mAb), the protein produced by such cells are in vogue. Such antibodies with single isotype have higher specificity, and the serological tests employed in show higher reproducibility compared to those with use of polyclonal antisera. There are several procedures of mAb production which vary considerably but the principle remains the same which states that antigens introduced into animals generally result in the stimulation of lymphocytes, some of which produce antibody of only one type, although the isotype may change. The developments in the field of cell culture and transfection technology have lead to the production of improved qualities of mAbs. In general, monoclonal antibodies are important reagents used in biomedical research, such as, in the field of diagnostics and therapeutics as well as targeted drug delivery systems. They have got importance not only for infectious diseases caused by microbes and parasites, but also for cancer, metabolic and hormonal disorders, in the diagnosis of lymphoid and myeloid malignancies and tissue typing, enzyme linked immunosorbent assay (ELISA) (especially blocking ELISA), radio immunoassay (RIA), serotyping of pathogens and their immunological intervention with passive antibody, anti-idiotype inhibition or magic bullet therapy with cytotoxic agents coupled with antimouse specific antibody. The application of mAbs in diagnosis of various livestock diseases is an important area of concern as these diseases are a major and increasingly important factor reducing livestock productivity in various parts of the world. In this context, the application of mAbs for diagnosis of important bacterial diseases viz., Anthrax, Brucellosis, Paratuberculosis, Leptospirosis, Listeriosis, Clostridial infections and Mycoplasmosis (CBPP), fungal diseases viz., Zygomycosis, Cryptococcosis, Histoplasmosis and Paracoccidiodomycosis, viral diseases viz., Foot-and-mouth disease (FMD), Infectious bovine rhinotracheitis/Infectious pustular vulvovaginitis (IBR/IPV), Rota viral diarrhoea, Blue tongue, Rabies, Classical swine fever and re-emerging viral diseases like Hendra and Nipah viral infections and parasitic diseases viz., dirofilariosis, and Trichinellosis and haemportozoan diseases (including Trypanosomiasis, Leishmaniasis, Anaplasmosis, infections caused by Plasmodium spp. as well as tick borne diseases) have been discussed thoroughly along with the specifications of the diagnostic assays for each disease for the convenience of the diagnosticians, researchers, scientists and students to employ such assays, both in field and laboratories to strengthen the disease control programme.

The diagnosis and differential diagnosis of granulocytic sarcoma

Objective： To discuss the diagnosis and differential diagnosis of granulocytic sarcoma （GS）. Methods： Six cases were reported in this paper. They were assessed by pathologists. Immunohistochemistry （IHC） stain and routine hematoxylin and eosin （H＆E） stain were applied. Results： All patients involved in different anatomic sites respectively including skin, lymph node, soft tissue, breast, cervix and penis. All cases were previously error diagnoses. Three of them were initially diagnosed as non-Hodgkin lymphoma （NHL）. One case of cervical lymph node lesion was first considered as metastasized carcinoma by clinician. One biopsied skin sample was initially reported as Karposi＇s sarcoma. And one breast case was suspicious of the Iobular carcinoma with the frozen samples without antecedent clinical history information. GS was accompanied with acute myeloid leukemia （AML） in one case and with acute lymphocytic leukemia （ALL） in one case. Histopathologically, blastic, immature and differentiated variants were found in four, one and one, respectively. Immunohistochemistry （IHC） showed that myeloperoxidase （MPO） and lysozyme were both found to be positive in all cases, CD43 was found in 5 of 6 cases. Three of six cases were CD68, CD15 and LCA positive. CD34 and CDl17 were positive in 1/5 and 1/6 cases, respectively. However, CD20 and CD3 were negative in all cases. Conclusion： GS was uncommon and it may be misdiagnosed easily in routine practice. Each area had its own character, but they had the common features too. It can be correctly diagnosed by combination of H＆E stain, IHC stain, peripheral blood and bone marrow. MPO and Lysozyme were necessary for the nature of granulocytes. In addition, CD43, CD68 and CD15 were very helpful.

Performance of a serological IgM and IgG qualitative test for COVID-19 diagnosis:An experimental study in Brazil

Qualitative antibody tests are an easy,point-of-care diagnostic method that is useful in diagnosing coronavirus disease 2019,especially in situations where reverse transcription-polymerase chain reaction is negative.However,some factors are able to affect its sensitivity and accuracy,which may contribute to these tests not being used as a first-line diagnostic tool.

基于黏膜sIgA抗体的非洲猪瘟病毒感染早期血清学检测方法的建立

目前的非洲猪瘟病毒(ASFV)血清学检测方法存在因采血引起交叉感染风险、抗体转阳滞后影响检测敏感性等问题,因此建立一种无创采样且可实现早期诊断的血清学方法对于在临床上ASFV感染的诊断与监测有重要意义。本研究以人工合成的方式构建了pFastbac1-P30-His重组表达载体,利用杆状病毒-昆虫细胞真核表达系统表达ASFV重组P30蛋白(SUMO-P30),用Ni柱亲和纯化后作为包被抗原,经过一系列条件优化,建立一种以猪口腔液中黏膜sIgA抗体为靶标的ASFV抗体间接ELISA方法。结果显示,真核重组表达的P30蛋白(SUMO-P30)约为47 ku,Western blot鉴定显示其具有良好的反应原性;经优化确定了ELISA最佳反应条件:包被量为1.0μg·mL^(-1),5%脱脂乳为最佳样品稀释液,与待检口腔液的最佳混合比例为2∶8,样品反应时间为120 min,鼠抗猪IgA-HRP最佳稀释度为1∶5000,反应时间为60 min,最佳显色时间为15 min。该方法检测ASFV感染口腔液抗体效价可达1∶32,与猪瘟病毒、猪伪狂犬病病毒、猪繁殖与呼吸综合征病毒黏膜抗体阳性口腔液均无交叉反应,具有良好的敏感性和特异性。利用该方法和ASFV商品化ELISA血清抗体检测试剂盒,分别检测感染ASFV强毒株或人工致弱株后同一头猪不同时间点的口腔液和血清样品,口腔液中黏膜sIgA抗体在感染后3~5 d S/P值就显著提升,而血清抗体在监测期内未检测到转阳。检测人工感染或同圈感染自然变异株后不同时间点的口腔液和血清样品,本研究建立的方法与商品化非洲猪瘟病毒血清抗体检测试剂盒检测的阳性符合率为100%,阴性符合率为37.5%,总符合率为61.5%。综上,本研究建立了一种无需采血,以口腔液为样品的ASFV黏膜sIgA抗体ELSIA检测方法,可实现ASFV感染的无创、早期、敏感诊断,为ASFV的监测和防控提供新的技术支持和补充。

类风湿性关节炎诊断中免疫学指标联合检测的应用价值分析

目的分析免疫学指标联合检测在类风湿性关节炎(RA)诊断中的应用价值。方法选择150例RA患者作为观察组,另选取同时段150例健康体检者作为对照组。两组均实施免疫学指标联合检测。对比两组类风湿因子(RF)、抗环瓜氨酸肽抗体(抗CCP抗体)、免疫球蛋白G(IgG)、补体C4和C3水平,抗CCP抗体、RF、抗角蛋白抗体(AKA)、抗核周因子抗体(APF)阳性率。结果观察组补体C4(0.19±0.07)g/L、补体C3(0.77±0.23)g/L低于对照组的(0.29±0.10)、(1.05±0.22)g/L,IgG(15.27±4.55)g/L高于对照组的(10.71±3.07)g/L(P<0.05)。观察组抗CCP抗体(302.58±98.48)IU/ml、RF(240.13±91.24)IU/ml均高于对照组的(6.57±1.23)、(7.25±2.34)IU/ml(P<0.05)。观察组抗CCP抗体阳性率82.67%(124/150)、RF阳性率81.33%(122/150)、AKA阳性率58.67%(88/150)、APF阳性率18.67%(28/150)均高于对照组的0、0.67%(1/150)、0、0(P<0.05)。结论临床在诊断RA中采取免疫学指标联合检测价值较高。