Isolation of Goose-origin scFv Antibodies Against Goose Parvovirus from Bacterial Display Antibody Libraries

Goose parvovirus(GPV)can cause a highly contagious and fatal gosling plague(GP)disease in goslings and muscoy ducklings.Here,three goose-origin neutralizing single chain variable fragment(scFv)antibodies against GPV SYG-61 were isolated.The genes of scFv antibodies were derived from goslings immunized with GPV SYG-61,and scFvs were subcloned into a pBSD vector for the construction of pBSD-scFv libraries.The pBSD-scFv libraries were screened following three rounds using VP2(protective antigen of GPV)as the bait by flow cytometry(FCM).After screening,the 15 clones with high mean fluorescence intensity(MFI)were isolated and sequenced.These 15 scFvs were expressed by pET-28a(+)in E.coli.The specificity and affinity of the 15 purified scFvs were successfully confirmed by ELISA.In the preliminary neutralization experiment on primary goose embryo fibroblast(GEF)in vitro,three of the 15 purified scFvs(named scFv-10,scFv-11 and scFv-50)showed significant neutralizing capacities.The study generated the first goose-origin neutralizing scFv against GPV and laid the foundation for the appearance of full-length goose-origin neutralizing monoclonal antibody against GPV.

Automatic Computer Analysis of Digital Images of Triple-Antibody-Stained Prostate Biopsies

Background: Worldwide, prostatic adenocarcinoma is the most common tumour type among men. Aim: The aim of the present investigation was to develop a computer program to identify normal prostate biopsies and distinguish them from biopsies showing premalignant alterations (LGPIN, HGPIN) and adenocarcinoma. Method: Prostate biopsies (n = 2094) taken from 191 consecutive men during 2016 were stained with triple immunehistochemisty (antibodies to AMACRA, p63 and CK 5). Digital images of the biopsies were obtained with a scanning microscope and used to develop an automatic computer program (CelldaTM), intended to identify the morphological alterations. Visual microscopic finding was used as a reference. Result: Of the 191 men, 121 (63.4%) were diagnosed as having prostate adenocarcinoma and 70 (36.6%) as having no malignancy on the basis of the visual microscopy. In comparison, computer analysis identified 134 (70.2%) men with malignant disease and 57 (29.8%) with non-malignant disease after exclusion of artifacts, which constituted 10.4% of areas (indicated as malignant disease). Discrepant results were recorded in 15 (7.9%) men, and in 14 of these cases, HGPIN and areas suggestive of early invasion were common. Thus, it was uncertain whether these cases should be regarded as malignant or not. The agreement between the visual examination and the computer analysis was 92.1% (kappa value 0.823, sensitivity 99.2 and specificity was 0.80). Conclusion: It seems that computer analysis could serve as an adjunct to simplify and shorten the diagnostic procedure, first of all by ensuring that normal prostate biopsies are sorted out from those sent for visual microscopic evaluation.

Analysis of Antisperm Antibody in Cervix Mucus for Azoospermic Men’s Wives

ELISA method was used to detect antisperm antibodies in cervix mucus. Out of 400 women whose husbands had sperms, 92 were antisperm antibody positive (23% ). 42 of 200 women with azoospermic partners were antisperm antibody positive (21% ). No difference was found between the two groups. Hence, antisperm antibodies in azoospermic men's wives may be caused by sperm'sadhering antigens in semina.

Methodological aspects of anti-human leukocyte antigen antibody analysis in solid organ transplantation

Donor human leukocyte antigen(HLA)-specific antibodies(DSA) play an important role in solid organ transplantation. Preexisting IgG isotype DSA are considered a risk factor for antibody mediated rejection, graft failure or graft loss. The post-transplant development of DSA depends on multiple factors including immunogenicity of mismatched antigens, HLA class Ⅱ typing of the recipient, cytokine gene polymorphisms, and cellular immunoregulatory mechanisms. De novo developed antibodies require special attention because not all DSA have equal clinical significance. Therefore, it is important for transplant clinicians and transplant immunologists to accurately characterize DSA. In this review, the contemporary immunological techniques for detection and characterization of anti-HLA antibodies and their pitfalls are described.

Numerical Analysis of Bacterial Meningitis Stochastic Delayed Epidemic Model through Computational Methods

Based on theWorld Health Organization(WHO),Meningitis is a severe infection of the meninges,the membranes covering the brain and spinal cord.It is a devastating disease and remains a significant public health challenge.This study investigates a bacterial meningitis model through deterministic and stochastic versions.Four-compartment population dynamics explain the concept,particularly the susceptible population,carrier,infected,and recovered.The model predicts the nonnegative equilibrium points and reproduction number,i.e.,the Meningitis-Free Equilibrium(MFE),and Meningitis-Existing Equilibrium(MEE).For the stochastic version of the existing deterministicmodel,the twomethodologies studied are transition probabilities and non-parametric perturbations.Also,positivity,boundedness,extinction,and disease persistence are studiedrigorouslywiththe helpofwell-known theorems.Standard and nonstandard techniques such as EulerMaruyama,stochastic Euler,stochastic Runge Kutta,and stochastic nonstandard finite difference in the sense of delay have been presented for computational analysis of the stochastic model.Unfortunately,standard methods fail to restore the biological properties of the model,so the stochastic nonstandard finite difference approximation is offered as an efficient,low-cost,and independent of time step size.In addition,the convergence,local,and global stability around the equilibria of the nonstandard computational method is studied by assuming the perturbation effect is zero.The simulations and comparison of the methods are presented to support the theoretical results and for the best visualization of results.

Cloning and Sequence Analysis of Light Variable Region Gene of Anti-human Retinoblastoma Monoclonal Antibody

Purpose: To clone the variable region gene of light chain of monoclonal antibody against human retinoblastoma and to analyze the characterization of its nucleotide sequence as well as amino acid sequence.Methods: Total RNA was extracted from 3C6 hybridoma cells secreting specific monoclonal antibody(McAb)against human retinoblastoma(RB), then transcripted reversely into cDNA with olig-dT primers.The variable region of the light chain (VL) gene fragments was amplified using polymeerase chain reaction(PCR) and further cloned into pGEM(R) -T Easy vector. Then, 3C6 VL cDNA was sequenced by Sanger's method.Homologous analysis was done by NCBI BLAST.Results: The complete nucleotide sequence of 3C6 VL cDNA consisted of 321 bp encoding 107 amino acid residues, containing four workframe regions(FRs)and three complementarity-determining regions (CDRs) as well as the typical structure of two cys residues. The sequence is most homological to a member of the Vk9 gene family, and its chain utilizes the Jkl gene segment.Conclusion: The light chain variable region gene of the McAb against human RB was amplified successfully , which belongs to the Vk9 gene family and utilizes Vk-Jk1 gene rearrangement. This study lays a good basis for constructing a recombinant antibody and for making a new targeted therapeutic agents against retinoblastoma.

Combatting Pathogenic Bacteria with Synthetic Immunotherapeutics from Chitosan:Antibody Recruiting at Engineered Bacterial Surface with Peptidoglycan Analogs

The emergence of microbial drug resistance,coupled with the paucity of new antibiotics poses an impending threat to public health.In this work,we drew inspiration from synthetic peptidoglycan oligomers and successfully constructed antibody-recruiting peptidoglycan analogs,2a-d,with excellent safety profiles and high efficiencies in recruiting antibodies across different conditions.Further,we demonstrated that these peptidoglycan analogs could be readily incorporated into bacteria cell walls,whereby both simple monoclonal and pooled human serum antibodies effectively congregated to the surfaceengineered bacteria,leading to the complete extermination of the engineered bacterial cells both in vitro and in vivo.Our peptidoglycan analog agents for recruiting endogenous antibodies to combat pathogenic bacteria enable further development of promising broad-spectrum immunotherapeutics.

Treatment‐related adverse events of antibody‐drug conjugates in clinical trials:A systematic review and meta‐analysis

Background:The wide use of antibody‐drug conjugates(ADCs)is transforming the cancer‐treatment landscape.Understanding the treatment‐related adverse events(AEs)of ADCs is crucial for their clinical application.We conducted a meta‐analysis to analyze the profile and incidence of AEs related to ADC use in the treatment of solid tumors and hematological malignancies.Methods:We searched the PubMed,Embase,and Cochrane Library databases for articles published from January 2001 to October 2022.The overall profile and incidence of all‐grade and grade≥3 treatment‐related AEs were the primary outcomes of the analysis.Results:A total of 138 trials involving 15,473 patients were included in this study.The overall incidence of any‐grade treatment‐related AEs was 100.0%(95%confidence interval[CI]:99.9%–100.0%;I2=89%)and the incidence of grade≥3 treatment‐related AEs was 6.2%(95%CI:3.0%–12.4%;I2=99%).Conclusions:This study provides a comprehensive overview of AEs related to ADCs used for cancer treatment.ADC use resulted in a high incidence of any‐grade AEs but a low incidence of grade≥3 AEs.The AE profiles and incidence differed according to cancer type,ADC type,and ADC components.

Preparation, Characterization, and Application of Antiharpin_(xoo) Antibody

Polyclonal antiharpinxoo rabbit antibody has been prepared successfully using purified harpinxoo protein as an immunogen. The ELISA titer of the antiserum against harpinxoo was about 1：2 000. Western blot analysis showed that the antiserum could bind to the expression harpinxoo protein in particular, hrfl, encoding harpinxoo, is an expression in transgenic rice, detected by antiharpinxoo rabbit antibody. The rabbit antibody against harpinxoo can be used to study further about the biological function, harpinxoo localization, and hrfl gene expression in other plants.

Development of Spectral Features for Monitoring Rice Bacterial Leaf Blight Disease Using Broad-Band Remote Sensing Systems

As an important rice disease, rice bacterial leaf blight (RBLB, caused by the bacterium Xanthomonas oryzae pv.oryzae), has become widespread in east China in recent years. Significant losses in rice yield occurred as a result ofthe disease’s epidemic, making it imperative to monitor RBLB at a large scale. With the development of remotesensing technology, the broad-band sensors equipped with red-edge channels over multiple spatial resolutionsoffer numerous available data for large-scale monitoring of rice diseases. However, RBLB is characterized by rapiddispersal under suitable conditions, making it difficult to track the disease at a regional scale with a single sensorin practice. Therefore, it is necessary to identify or construct features that are effective across different sensors formonitoring RBLB. To achieve this goal, the spectral response of RBLB was first analyzed based on the canopyhyperspectral data. Using the relative spectral response (RSR) functions of four representative satellite or UAVsensors (i.e., Sentinel-2, GF-6, Planet, and Rededge-M) and the hyperspectral data, the corresponding broad-bandspectral data was simulated. According to a thorough band combination and sensitivity analysis, two novel spectralindices for monitoring RBLB that can be effective across multiple sensors (i.e., RBBRI and RBBDI) weredeveloped. An optimal feature set that includes the two novel indices and a classical vegetation index was formed.The capability of such a feature set in monitoring RBLB was assessed via FLDA and SVM algorithms. The resultdemonstrated that both constructed novel indices exhibited high sensitivity to the disease across multiple sensors.Meanwhile, the feature set yielded an overall accuracy above 90% for all sensors, which indicates its cross-sensorgenerality in monitoring RBLB. The outcome of this research permits disease monitoring with different remotesensing data over a large scale.

Analysis of clinical features and prognosis of anti-glomerular basement membrane antibody positive patients with anti-neutrophil cytoplasmic antibodies

Objective To investigate the characteristics and outcome of glomerulonephritis in patients with both antineutrophil cytoplasmic antibody and anti-glomerular basement membrane antibody.Methods The sera of 23 antiGBM glomerulonephritis patients were collected and were tested for ANCA respectively.Characteristics and outcome of patients with coexisting anti-GBM antibody

Semen analysis in chronic bacterial prostatitis： diagnostic and therapeutic implications

The significance and diagnostic value of semen analysis in chronic bacterial prostatitis has been extensively debated and remains controversial. To investigate the diagnostic relevance of semen culture in the bacteriological workup of prostatitis patients, we retrospectively analyzed a clinical database of 696 symptomatic patients. All patients were routinely subjected to a four-glass test, followed by semen culture and analysis. This allowed to dissect from the database three different diagnostic scenarios, and to compare the ＇two-glass＇ pre-/post- massage test and the standard ＇four-glass＇ test with a ＇five-glass＇ test （four-glass plus post-VB3 semen culture）. The ＇five-glass＇ test showed 3.6- or 6.5-fold increases in relative sensitivity and lesser reductions （-13.2% or -14.7%） in relative specificity for traditional uropathogens （TUs） compared with the four-glass or two-glass test, respectively. The area under the ROC curve and Jouden＇s index were increased, whereas positive and negative likelihood ratios were lower than comparators, indicating that the ＇five-glass＇ assay may be superior in confirming the negative outcome of both standard tests. The five-, four-, and two-glass tests detected TUs （Enterobacteriaceae, Enterococci, etc.） in 120, 33, and 20 patients and unusual pathogens （Streptococci, other Gram-positive species, Mycoplasmata, and others） in 130, 56, and 45 patients, respectively. When patients were subjected to pharmacological treatment, including a combination of a fluoroquinolone and a macrolide, no differences in eradication rates were observed between groups diagnosed with different tests, irrespective of pathogen category. Eradication was associated with long-term sign/symptom remission; no significant intergroup differences in sign/symptom scores were observed throughout a 24-month off-therapy follow-up period. In conclusion, our data support the usefulness of semen analysis in the diagnostic workup ofprostatitis patients when this test is used to complement the four-glass Meares and Stamey test. Improvement of microbiological assays conveys important diagnostic and therapeutic implications.

The efficacy and safety of besifloxacin for acute bacterial conjunctivitis: a Meta-analysis

AIM: To evaluate the relative efficacy and safety of besifloxacin for treatment of acute bacterial conjunctivitis.METHODS: A comprehensive search in PubMed, EMBASE Web of Science, Cochrane Central Database and CNKI was undertaken for randomized controlled trials(RCTs) comparing besifloxacin with other treatments or placebo. The primary outcome measures were clinical resolution, rates of bacterial eradication, individual clinical outcomes, cure rates, and bacterial eradication rates of different kinds of pathogens. Safety outcomes were the number of adverse effects(AEs). The final search was performed on August 2018.RESULTS: Eight RCTs were included. Five studies compared the efficacy and safety of besifloxacin with placebo, 2 studies compared besifloxacin with moxifloxacin, and 1 study compared besifloxacin with gatifloxacin. A total of 3105 patients met the inclusion criteria. Besifloxacin presented higher efficacy and safety than did placebo in clinical resolution, rates of bacterial eradication, individual clinical outcomes, cure rates, bacterial eradication rates of different kinds of pathogens and the number of AEs. There was no significant difference between besifloxacin and moxifloxacin or gatifloxacin in the comparison items mentioned above.CONCLUSION: Besifloxacin is highly effective and safe for treatment of acute bacterial conjunctivitis. Further comparative trials regarding the effect of besifloxacin for treatment of acute bacterial conjunctivitis will aid in treatment decisions.

Treating with besifloxacin for acute bacterial conjunctivitis: a Meta-analysis

AIM: To evaluate the relative efficacy and safety of besifloxacin for treatment of acute bacterial conjunctivitis. METHODS: A comprehensive search in PubM ed, EMBASE Web of Science, Cochrane Central Database and CNKI was undertaken for randomized controlled trials(RCTs) comparing besifloxacin with other treatments or placebo. The primary outcome measures were clinical resolution, rates of bacterial eradication, individual clinical outcomes, cure rates, and bacterial eradication rates of different kinds of pathogens. Safety outcomes were the number of adverse effects(AEs). The final search was performed on August 2018. RESULTS: Six RCTs were included. Four studies compared the efficacy and safety of besifloxacin with placebo, 1 study compared besifloxacin with moxifloxacin, and 1 study compared besifloxacin with gatifloxacin. A total of 2780 patients met the inclusion criteria. Besifloxacin presented higher efficacy and safety than did placebo in clinical resolution, rates of bacterial eradication, individual clinical outcomes, cure rates, bacterial eradication rates of different kinds of pathogens and the number of AEs. There was no significant difference between besifloxacin and moxifloxacin or gatifloxacin in the comparison items mentioned above. CONCLUSION: Besifloxacin is highly effective and safe for treatment of acute bacterial conjunctivitis. Further comparative trials regarding the effect of besifloxacin for treatment of acute bacterial conjunctivitis will aid in treatment decisions.

Analysis of Bacterial Communities Present in Agaricus bisporus Phase Ⅱ Compost Using Amplified Ribosomal DNA Restriction Analysis(ARDRA)(Ⅱ)——Analysis of Bacterial Communities

Changes in the bacterial communities inhabiting Agaricus bisporus compost during Phase Ⅱ treatment were monitored using amplified rDNA restriction analysis(ARDRA).Sequence analysis data indicated that the bacterial communities associated with the compost samples were far richer in composition when determined by ARDRA compared with traditional methods based on bacterial isolation.Furthermore,the composition of the communities,especially in terms of the dominant bacteria during each of the four treatment stages(beginning of Phase Ⅱ,after pasteurization,5 days aeration,end of Phase Ⅱ,respectively),were very different.Restriction endonuclease digestion of mainly bacterial clones from four 16S rDNA libraries,C1,C3,C6 and C7,previously constructed on the basis of 16S rDNA fragments amplified from the four different stages of the Phase Ⅱ composting process,revealed the presence of unique microbial types.Thermophilic bacteria belonging to the Bacilli,and previously unreported strains of Trichococcus,Planococcus,Caryophanon,and subclass γ-Proteobacteria,were identified among the clones from the compost sampled during the aeration period(C6).Bacteria belonging to Thermus thermophilus and subclass α-Proteobacteria were detected in C1 and C7 composts sampled at the beginning and the end of Phase Ⅱ,respectively.Clones of some uncultured bacteria were also scored.

ANALYSIS ON EPITOPES OF IGM WITH MONOCLONAL ANTI-ISOTYPIC AND ANTI-IDIOTYPIC ANTIBODIES AGAINST IgM FROM B-CLL

A double antibodies additivity ELISA test was employed to identify the epltopes which can be recognized by monoclonal antibodies (McAbs) against IgM from B chronic lymphocyte leukemia (B-CLL). The computer grouping programme analysis showed that 4 and- isotypic MaAbs could be divided into two groups and 10 anti- idiotype McAbs could be divided into four groups. The result was consistent with that of the indirect sandwich ELISA and inhibition ELISA test. It suggested that there were at least 6 distinct IgM epitopes which can react specifically with 14 McAbs. Our study indicated that the combination of the additivity ELISA test and the computer grouping programme analysis is of help in studying the relationship of the structure and function of antigen.

Correlation between dominant bacterial community and non-volatile organic compounds during the fermentation of shrimp sauces

Shrimp sauce,one of the traditional salt-fermented food in China,has a unique flavor that is influenced by the resident microflora.The quality of salt-fermented shrimp sauce was evaluated in this work by determining the total volatile basic nitrogen(TVB-N),the amino acid nitrogen(AAN),organic acid,5’-nucleotide and free amino acids(FAA).Moreover,the dynamics of microbial diversity during processing was investigated by using high-throughput sequencing technology.The results showed that the AAN,TVB-N,organic acid,5’-nucleotide and FAA content were in range of 0.93-1.42 g/100 mL,49.91-236.27 mg/100 mL,6.65-20.68 mg/mL,3.51-6.56 mg/mL and 81.27-102.90 mg/mL.Among the microbial diversity found in the shrimp sauce,Tetragenococcus,Flavobacterium,Polaribacter,Haematospirillum and Staphylococcus were the predominant genera.Correlation analysis indicated that the bacteria Tetragenococcus and Staphylococcus were important in the formation of non-volatile compounds.Tetragenococcus positively correlated with a variety of FAAs;Staphylococcus positively correlated with 5’-nucleotides.The analysis indicated that Tetragenococcus and Staphylococcus were the core genera affecting non-volatile components.These findings indicate the dynamics of the bacterial community and non-volatile components inter-relationships during shrimp sauce fermentation and provide a theoretical basis for improving the fermentation process of shrimp sauce.

Comparing different domains of analysis for the characterisation of N-glycans on monoclonal antibodies

With the size of the biopharmaceutical market exponentially increasing,there is an aligned growth in the importance of data-rich analyses,not only to assess drug product safety but also to assist drug development driven by the deeper understanding of structure/function relationships.In monoclonal antibodies,many functions are regulated by N-glycans present in the constant region of the heavy chains and their mechanisms of action are not completely known.The importance of their function focuses analytical research efforts on the development of robust,accurate and fast methods to support drug development and quality control.Released N-glycan analysis is considered as the gold standard for glycosylation characterisation;however,it is not the only method for quantitative analysis of glycoform heterogeneity.In this study,ten different analytical workflows for N-glycan analysis were compared using four monoclonal antibodies.While observing good comparability between the quantitative results generated,it was possible to appreciate the advantages and disadvantages of each technique and to summarise all the observations to guide the choice of the most appropriate analytical workflow according to application and the desired depth of data generated.

Role of lumbar puncture in clinical outcome of suspected acute bacterial meningitis

BACKGROUND Meningitis remains a significant source of mortality and morbidity,with an incidence of 1 per 100000 persons in the United States.Guidelines recommend obtaining blood cultures and cerebrospinal fluid(CSF)studies in patients presenting with acute meningitis syndrome,and beginning treatment with broad spectrum antibiotics based on the age and certain predisposing conditions.In some patients however,the diagnostic lumbar puncture(LP)is not performed due to a multitude of reasons,ranging from increased intracranial pressure to failed attempt.In such situations,appropriate therapy is initiated empirically and often continued without establishment of a definitive diagnosis.AIM To determine whether a diagnostic LP in acute meningitis syndrome was associated with a better outcome and less duration of antibiotic therapy,along with potential causes for deferral of procedure.METHODS A retrospective study was conducted amongst the patients presenting to a 360 bedded community hospital in central Massachusetts with a diagnosis of acute meningitis syndrome between January 2010-September 2022.The electronic health records were accessed to collect necessary demographic and clinical data,including etiology of meningitis,lumbar puncture results,reason for procedure deferral,duration of antibiotic therapy and clinical outcome.The patients were subsequently divided into two groups based on whether they received a LP or not,and data was analyzed.RESULTS A total of 169 patients admitted with acute meningitis syndrome between September 2010-2022 were included in the study.The mean age of the participants was 54.3 years(SD+/-19.2 years).LP was performed for 130(76.9%)participants,out of which,28(21.5%)showed some growth in CSF culture.The most commonly identified organism was streptococcus pneumoniae.Amongst the 39 patients in whom LP was deferred,the major reasons recorded were:Body habitus(n=6,15.4%),and unsuccessful attempt(n=4,10.3%).While 93(71.5%)patients with LP received antibiotic therapy,only 19(48.7%)patients without LP received the antibiotics,with the principal reason being spontaneous improvement in sensorium without any diagnosed source of infection.The mean duration of antibiotic use was 12.3 days(SD+/-5.6)in the LP group and 11.5 days(SD+/-7.0)in the non-LP group(P=0.56;statistically not significant).We observed higher long term sequalae in the non-LP group(n=6,15.4%)compared to the LP group(n=9,6.9%).Similarly,the death rate was higher in the non-LP group(n=7,18.0%)compared to the LP group(n=9,6.9%).CONCLUSION LP remains the cornerstone for diagnosing meningitis,but often CSF results are unavailable,leading to empiric treatment.Our study identified that body habitus and unsuccessful attempts were the most common reasons for LP not being performed,leading to empiric antibiotic coverage.There was no difference between the duration of antibiotics received by the two groups,but a lower proportion of patients without LP received antibiotics,attributed to a spontaneous improvement in sensorium.However,the residual neurological sequelae and death rates were higher in patients without LP,signifying a potential under-treatment.A LP remains crucial to diagnose meningitis,and a lack of CSF analysis predisposes to under-treatment,leading to higher neurological sequelae and increased chances of death.