Anti-PD1 antibody and not anti-LAG-3 antibody improves the antitumor effect of photodynamic therapy for treating metastatic breast cancer

Photodynamic therapy(PDT)has limited effects in treating metastatic breast cancer.Immune checkpoints can deplete the function of immune cells;however,the expression of immune checkpoints after PDT is unclear.This study investigates whether the limited e±cacy of PDT is due to upregulated immune checkpoints and tries to combine the PDT and immune checkpoint inhibitor to observe the e±cacy.A metastatic breast cancer model was treated by PDT mediated by hematoporphyrin derivatives(HpD-PDT).The anti-tumor effect of HpD-PDT was observed,as well as CD4þT,CD8þT and calreticulin(CRT)by immunohistochemistry and immunofluorescence.Immune checkpoints on T cells were analyzed byflow cytometry after HpD-PDT.When combining PDT with immune checkpoint inhibitors,the antitumor effect and immune effect were assessed.For HpD-PDT at 100 mW/cm2 and 40,60 and 80 J/cm2,primary tumors were suppressed and CD4þT,CD8þT and CRT were elevated;however,distant tumors couldn't be inhibited and survival could not be prolonged.Immune checkpoints on T cells,especially PD1 and LAG-3 after HpD-PDT,were upregulated,which may explain the reason for the limited HpD-PDT effect.After PDT combined with anti-PD1 antibody,but not with anti-LAG-3 antibody,both the primary and distant tumors were signi-cantly inhibited and the survival time was prolonged,additionally,CD4þT,CD8þT,IFN-þCD4þT and TNF-þCD4þT cells were signi-cantly increased compared with HpD-PDT.HpD-PDT could not combat metastatic breast cancer.PD1 and LAG-3 were upregulated after HpD-PDT.Anti-PD1 antibody,but not anti-LAG-3 antibody,could augment the antitumor effect of HpD-PDT for treating metastatic breast cancer.

Monoclonal antibodies as therapeutic agents in oncology and antibody gene therapy

Antibodies as therapeutic agents are mostly used in oncology, as illustrated by their applications in lymphoma, breast cancer or colorectal cancer. This review provides a brief historical sketch of the development of monoclonal antibodies for cancer treatment and summarizes the most significant clinical data for the best-established reagents to date. It also discusses strategies to improve the anti-tumor efficacy of antibody therapy, including antibody gene therapy and exploitation of bone marrow derived primary mesenchymal stem cells as the antibody gene transporter.

Preparation of Polyclonal Antibodies Against Testis-specific Protease 50 and Characterization of Antibody Specificity

Testis-specific protease 50 （TSP50） is a testis-specific oncogene, which is abnormally activated in most tested patients with breast cancer. This property makes it an attractive molecular marker and a promising target for the diagnosis and therapy of breast cancer. In order to obtain the protective and specific polyclonal antibodies for further research, TSPS0 cDNA was amplified by RT-PCR from normal human testicular tissue, and inserted into eukaryotic expression vector PeDNA3.1. Rabbit anti-TSPS0 polyclonal antibodies were prepared by means of intramuscular injection of peDNA3.1-TSPS0 into the rabbits. Titem of the anti-sera were measured by ELISA and Western blotting with the E. coli cell lysate containing the induced GST-TSPS0 fusion protein as an antigen. In addition, we examined the expression of TSPS0 in both breast cancer cell line MCF-7 and breast cancer tissue by immunofluorescent and immunohistochemistry analysis.

PRODUCTION AND APPLICATION OF MONOCLONAL ANTIBODY TO POLYAMINE (PREPARATION AND CHARACTERISTICS OF MONOCLONAL ANTIBODIES AGAINST SPERMIDINE)

A monoclonal antibody was first prepared by fusion of mouse myeloma cells (SP2/0-Ag-14) with spleen cells isolated from male BALB/ c mice immunized with spermidine-bovine serum albumin conjugate (SPD- BSA). The hybridoma cell line producing antibody specific for spermidine was cultured in vitro and after i. p. into mice, the ascitic fluid gave suitably high dilution titres (1: 106) by enzyme immunoassay. This monoclonal antibody is of IgG1 class and the bimolecular compleex with molecular weight of 52KD and 27 KD. The monoclonal antibody was clearly specific to spermidine comparing with spermine or putriscine. Monclonal antibody may prove to be useful in the rapid diagnosis and evaluation of patients with cancer.

ISOLATION OF ENDOTOXIN-SPECIFIC ANTIBODIES BY SELECTION OF AN SINGLE CHAIN PHAGE ANTIBODY LIBRARY

Objective: To isolate murine anti endotoxin single chain phage antibody from a constructed library. Methods: Total RNA was firstly extracted from murine splenic cells and mRNA was reverse-transcribed into cDNA. Then the designed primers were used to amplify the variable region genes of the heavy and light chain (VH, VL) with polymerase chain reaction. The linker was used to assemble the VH and VL into ScFv, and the NotI and SfiI restriction enzymes were used to digest the ScFv in order to ligate into the pCANTAB5E phagemid vector that was already digested with the same restriction enzymes. The ligated vector was then introduced into competent E.coli TG1 cells to construct a single-chain phage antibody library. After rescued with M13KO7 helper phage, recombinant phages displaying ScFv fragments were harvested from the supernatant and selected with endotoxin. The enriched positive clones were reinfected into TG1 cells. Finally, 190 clones were randomly selected to detect the anti endotoxin antibody with indirect ELISA. Results: The titer of anti endotoxin in murine sera was 1:12,800. The concentration of total RNA was 12.38 μg/ml. 1.9×107 clones were obtained after transformed into TG1. 3×104 colonies were gotten after one round panning. Two positive colonies were confirmed with indirect ELISA among 190 randomly selected colonies. Conclusion: A 1.9×107 murine anti endotoxin single chain phage antibody library was successfully constructed. Two anti endotoxin antibodies were obtained from the library.

Analysis of Specific Th1/Th2 Helper Cell Responses and IgG Subtype Antibodies in Anti-CD4 Monoclonal Antibody Treated Mice with Autoimmune Cardiomyopathy

The cytokine repertoire of ADP/ATP carrier-specific humoral immune responses and the cytokine-dependent anti-ADP/ATP carrier antibody IgG subclasses were examined in a cohort of ADP/ATP carrier-immunized BALB/c mice treated with anti-CD4 monoclonal antibody. Eighteen male BALB/c mice （6–8 weeks old） were randomized into 3 groups： dilated cardiomyopathy （DCM） group, DCM-tolerance （Tol） group and control group. The mice in DCM group were immunized with the peptides derived from human ADP/ATP carrier protein for 6 months and mice in the control group were sham-immunized, while the mice in DCM-Tol group were immunized with ADP/ATP carrier protein and anti-CD4 McAb simultaneously. Serum autoantibody against ADP/ATP carrier and IgG subclasses were measured by ELISA, intracellular cytokines IFN-γ and IL-4 of Th cells were moni- tored with flow cytometry, and splenic T cell cytokines IFN-γ, IL-2, IL-4 and IL-6 were detected by using real-time fluorescent quantitative PCR. The results showed that the autoantibody against ADP/ATP carrier was found in all mice in DCM group, and the antibody level, serum IgG1 and IgG2a subclasses, cytokines in T cells and Th cells were all elevated in DCM group, as compared with those in control group （P〈0.01）. On the other hand, in DCM-Tol group, the autoantibody level and contents of all the cytokines were significantly different from those in DCM group （P〈0.01）, and were close to those in control group. And the levels of IgG1, IgG2a, IgG2b and IgG3 were influenced, to varying degrees, by anti-CD4 McAb as compared with those in DCM group. All these four types of IgG subclasses were substantially decreased in DCM-Tol group as compared with DCM group. It is concluded that the treatment with anti-CD4 McAb could prevent the activation of T cells, reverse the abnormal secretion of cytokines and the imbalance between Th1/Th2 cell subsets and abnormal production of autoantibody against ADP/ATP carrier, and eventually avoid myocardial injuries.

Autoantibody profiles in autoimmune hepatitis and chronic hepatitis C identifies similarities in patients with severe disease

To determine how the auto-antibodies (Abs) profiles overlap in chronic hepatitis C infection (CHC) and autoimmune hepatitis (AIH) and correlate to liver disease.METHODSLevels of antinuclear Ab, smooth muscle antibody (SMA) and liver/kidney microsomal-1 (LKM-1) Ab and markers of liver damage were determined in the sera of 50 patients with CHC infection, 20 AIH patients and 20 healthy controls using enzyme linked immunosorbent assay and other immune assays.RESULTSWe found that AIH patients had more severe liver disease as determined by elevation of total IgG, alkaline phosphatase, total serum bilirubin and serum transaminases and significantly higher prevalence of the three non-organ-specific autoantibodies (auto-Abs) than CHC patients. Antinuclear Ab, SMA and LKM-1 Ab were also present in 36% of CHC patients and related to disease severity. CHC cases positive for auto-Abs were directly comparable to AIH in respect of most markers of liver damage and total IgG. These cases had longer disease duration compared with auto-Ab negative cases, but there was no difference in gender, age or viral load. KLM-1+ Ab CHC cases showed best overlap with AIH.CONCLUSIONAuto-Ab levels in CHC may be important markers of disease severity and positive cases have a disease similar to AIH. Auto-Abs might have a pathogenic role as indicated by elevated markers of liver damage. Future studies will unravel any novel associations between these two diseases, whether genetic or other.

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.

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.

THE USE OF ANTI-HUMAN GLIOMA MONOCLONAL ANTIBODIES FOR TARGETING CHEMOTHERAPY OF BRAIN GLIOMAS(PREPARATION AND CYTOTOXIC PROPERTIES OF ANTIBODY-ADRIAMYCIN IMMUNOCONJUGATES)

Immunoconjugates are antibody-drug hybrid molecules which combine the exquisite selectivity or monoclonal antibodies with the potent toxicity of anticancer agents. A monoclonal antibody SZ39 against human brain gliomas was used as a drug carrier. Adriamycin (ADR) was bound covalently to SZ39 to form a SZ39-ADR conjugate. The cytotoxic activity of the SZ39-ADR conjugate was tested in vitro and demonstrated potent and specific killing of cells derived from a human malignant glioma. 50% inhibitory concentration (IC50) for SZ39-ADR to 'target' cells was 8.14×10-9 M. An index of specificity between 'target' and 'non-target' cells was calculated to be 88-fold. These data suggest that the SZ39-ADR may use as a potent and cell type-specific agent and is a likely candidate for the targeting chemotherapy of malignant gliotnas.

Preparation of anti-canine interleukin-31 receptor alpha polyclonal antibody and evaluation of its therapeutic efect in canine atopic dermatitis

Canine atopic dermatitis(CAD)is a prevalent genetically susceptible infammatory and pruritic allergic skin condition afecting not only the health of dogs but also the quality of life of their owners.Interleukin-31(IL-31)and interleukin-31 receptor alpha(IL-31RA)are essential for the development of pruritus in primates and mice.Hence,it is expected that inhibiting IL-31RA will be an efective approach to alleviate pruritus.The purpose of the study was to produce anti-canine IL-31RA polyclonal antibodies(anti-IL-31RA pAbs)and evaluate their efcacy in inhibiting house dust mite(HDM)-evoked pruritic responses.Dogs were immunized with antigens formed by IL-31RA recombinant short peptides coupled to BSA to produce anti-IL-31RA pAbs.The CAD model was developed by using HDM allergen stimulation,and the efects of IL-31RA pAbs on the reduction of pruritus in CAD model dogs were examined.The Canine Atopic Dermatitis Extent and Severity Index(CADESI)-4 and pruritus Visual Analog Scale(pVAS)were utilized to evaluate pruritic responses,and skin tissue samples were collected from the inguinal area for pathological assessment of skin infammatory cell infltration.The results showed that anti-IL-31RA pAbs with high titers(1:128,000)and specifcity were efectively produced.In the CAD model group,the severity of skin damage,pruritus score,infammatory cell infltration and level of infammatory factors were considerably elevated.Anti-IL-31RA pAbs relieved pruritic behavior and dermatitis in dogs compared to placebo-treated dogs.In conclusion,anti-IL-31RA pAbs efectively suppressed CAD in vivo and are anticipated to be an efective novel treatment for pruritic skin disorders such as CAD.

Anti-OX40 Antibody Combined with HBc VLPs Delays Tumor Growth in a Mouse Colon Cancer Model

Objective Combination immunotherapy strategies targeting OX40,a co-stimulatory molecule that can enhance antitumor immunity by modulating the proliferation,differentiation,and effector function of tumor-infiltrating T cells,have attracted much attention for their excellent therapeutic effects.In this study,we aimed to evaluate the antitumor efficacy of combined anti-OX40 and hepatitis B core viruslike particles(HBc VLPs)therapy using a mouse colon cancer model.Methods Humanized B-h OX40 mice were injected subcutaneously with MC38 colon tumor cells and treated with HBc VLPs+anti-h OX40 antibody.Tumor growth was monitored.Flow cytometric analysis was performed to evaluate the populations of T cell subsets in the tumors.Results The combination of anti-OX40 with HBc VLPs resulted in a significant delay in tumor growth,suggesting that a potent antitumor immunity was induced by the combination therapy.Further studies revealed that HBc VLPs+anti-OX40 treatment induced a significant increase in effector T cells(Teffs)and a significant decrease in regulatory T cells(Tregs)in the tumor microenvironment(TME),which accounted for the synergistic antitumor effect of anti-OX40 in combination with HBc VLPs.Conclusion Combination therapy of anti-h OX40 and HBc VLPs provides synergistic antitumor activity in colon cancer-bearing mice,which may represent a potential design strategy for cancer immunotherapy.

Antibody Levels and Infection Status of Pertussis in the Population under Pertussis Resurgence in Guangxi in 2018:A Cross-Sectional Survey

Objective Pertussis cases have increased markedly since 2018 in Guangxi.The aim of this study was to evaluate antibody levels and the infection status of pertussis in the resident population.Method A total of 10,215 serum samples from residents were collected from August-November 2018 and tested for anti-pertussis IgG and toxin IgG using the enzyme-linked immunosorbent assay(ELISA).Results Of the collected samples,1,833(17.94%)tested positive for anti-pertussis IgG,with the median concentration of 16.06 IU/mL.Antibody level<10 IU/mL accounted for more than 60%in children under 4 years of age,but declined with age,whereas the percentages of the other three levels(10-40,40-50,and≥50 IU/mL)increased almost with age(P<0.001).Moreover,7,924 samples were selected for anti-pertussis toxin IgG,of which 653(8.24%)tested positive(≥40 IU/mL)with the median concentration of 5.89 IU/mL,and 204 participants(2.56%)had recent pertussis infection(≥100 IU/mL).Among the different age groups,the highest rates of positivity and recent infection were observed at 11-20 years of age,the lowest positivity rate at 5 years of age,and the lowest recent infection rate at 4 years of age(P<0.001,P=0.005,respectively).Conclusion The survey results showed that all age groups in Guangxi lacked immunity against pertussis,which was one of the main factors contributing to the resurgence of pertussis in 2018.In addition,the prevalence of pertussis is relatively high in Guangxi,and its incidence is seriously underestimated,especially in adolescents and adults.

Antibody-platinum(IV)prodrugs conjugates for targeted treatment of cutaneous squamous cell carcinoma

Antibody-drug conjugates(ADCs)are a new type of targeting antibodies that conjugate with highly toxic anticancer drugs via chemical linkers to exert high specificity and efficient killing of tumor cells,thereby attracting considerable attention in precise oncology therapy.Cetuximab(Cet)is a typical antibody that offers the benefits of good targeting and safety for individuals with advanced and inoperable cutaneous squamous cell carcinoma(cSCC);however,its anti-tumor activity is limited to a single use.Cisplatin(CisPt)shows good curative effects;however,its adverse effects and non-tumor-targeting ability are major drawbacks.In this study,we designed and developed a new ADC based on a new cytotoxic platinum(IV)prodrug(C8Pt(IV))and Cet.The so-called antibody-platinum(IV)prodrugs conjugates,named Cet-C8Pt(IV),showed excellent tumor targeting in cSCC.Specifically,it accurately delivered C8Pt(IV)into tumor cells to exert the combined anti-tumor effect of Cet and CisPt.Herein,metabolomic analysis showed that Cet-C8Pt(IV)promoted cellular apoptosis and increased DNA damage in cSCC cells by affecting the vitamin B6 metabolic pathway in tumor cells,thereby further enhancing the tumor-killing ability and providing a new strategy for clinical cancer treatment using antibody-platinum(IV)prodrugs conjugates.

Antibody Titer of Newcastle Disease in Vaccinated and Non-vaccinated Local Chicken of Cambodia

The experiment was conducted at Veterinary Research Station of Faculty of Veterinary Medicine,Royal University of Agriculture.The experimental period lasted 60 days,starting from October 1st to November 30th 2022.CRD(Completely Randomized Design)was used with 2 treatments/groups,vaccination group and non-vaccination group“control”,and 6 replications.The vaccination groups received two times of vaccination by dropping into the ocular at 7 days and 21 days.Meanwhile,blood samples were collected 3 times to detect the antibody level of ND(Newcastle Disease)and contained 21 days old,35 days old and 49 days old chicks.The ELISA(Enzyme-Linked Immunosorbent Assay)was performed to detect the antibody of those 2 groups.The result of finding showed that the S/P(Sample to Positive)ratio of control at 21 days,was very low,even in 3rd quartile,which was below the threshold.However,the vaccination group was relatively high,even in 1st quartile,which was higher than the threshold.At 35 days,S/P ratio of control group was still very low,but a bit higher than at 21 days.Meanwhile,the vaccination group was still high,even in 1st quartile,and two-time higher than at 21 days,but an increasing number of samples developed less antibody than threshold,accounting for 12.22%.At 49 days,the control group was still very low,even in 3rd quartile,but a bit higher than at 21 days and 35 days,and was close to the threshold.The vaccination group was still relatively high,even in 1st quartile but lower than three times comparing to 35 days.However,in this age,the number of chickens that developed antibody seemed to be increased in the control group,vice versa for vaccination group.The average S/P ratio was different significant(p<0.001),where vaccination had higher S/P ratio than control.It was similar finding for log-titer,the vaccination had higher figure(p<0.001).The risk of infection of ND was higher in control group,but it will reduce by increasing the age of chicken,while vaccination group was decreased by increasing age,especially at 49 days and we need to consider another vaccination to get full protection.

Quantitative hepatitis B core antibody and quantitative hepatitis B surface antigen:Novel viral biomarkers for chronic hepatitis B management

The management of hepatitis B virus(HBV)infection now involves regular and appropriate monitoring of viral activity,disease progression,and treatment response.Traditional HBV infection biomarkers are limited in their ability to predict clinical outcomes or therapeutic effectiveness.Quantitation of HBV core antibodies(qAnti-HBc)is a novel non-invasive biomarker that may help with a variety of diagnostic issues.It was shown to correlate strongly with infection stages,hepatic inflammation and fibrosis,chronic infection exacerbations,and the presence of occult infection.Furthermore,qAnti-HBc levels were shown to be predictive of spontaneous or treatment-induced HBeAg and HBsAg seroclearance,relapse after medication termination,re-infection following liver transplantation,and viral reactivation in the presence of immunosuppression.qAnti-HBc,on the other hand,cannot be relied on as a single diagnostic test to address all problems,and its diagnostic and prognostic potential may be greatly increased when paired with qHBsAg.Commercial qAnti-HBc diagnostic kits are currently not widely available.Because many methodologies are only semi-quantitative,comparing data from various studies and defining universal cut-off values remains difficult.This review focuses on the clinical utility of qAnti-HBc and qHBsAg in chronic hepatitis B management.

A Comparative Study on Detection of the Expression of Alternative Oxidase in Tobacco Callus with the Monoclonal Antibody Against Alternative Oxidase and Antibody Against-Synthetic Polypeptide Antibody

从经过不同温度处理的烟草 (NicotianarusticaL .)愈伤组织中提取并纯化线粒体蛋白 ,分别与交替氧化酶的单克隆抗体和抗合成多肽抗体进行免疫杂交。结果表明 :交替氧化酶的含量随温度的下降而显著上升 ;单克隆抗体的特异性较高于抗合成多肽抗体 ,但后者与交替氧化酶同样有良好的亲和性。因此 ,用合成多肽方法制备的抗体可以用于交替氧化酶的研究中。

Strategies to enhance monoclonal antibody uptake and distribution in solid tumors

Despite the significant resources dedicated to the development of monoclonal antibody(m Ab)therapies for solid tumors,the clinical success,thus far,has been modest.Limited efficacy of m Ab in solid tumors likely relates to unique aspects of tumor physiology.Solid tumors have an aberrant vasculature and a dense extracellular matrix that slow both the convective and diffusive transport of m Abs into and within tumors.For m Abs that are directed against cellular antigens,high antigen expression and rapid antigen turnover can result in perivascular cells binding to and eliminating a significant amount of extravasated m Ab,limiting m Ab distribution to portions of the tumor that are distant from functional vessels.Many preclinical investigations have reported strategies to improve m Ab uptake and distribution;however,to our knowledge,none have translated into the clinic.Here,we provide an overview of several barriers in solid tumors that limit m Ab uptake and distribution and discuss approaches that have been utilized to overcome these barriers in preclinical studies.

Construction of human Fab library and screening of a single-domain antibody of amyloid-beta 42 oligomers

Screening humanized antibodies from a human Fab phage display library is an effective and quick method to obtain beta-amyloid oligomers. Thus, the present study prepared amyloid-beta 42 oli- gomers and constructed a have human Fab phage display library based on blood samples from six healthy people. After three rounds of biopanning in vitro, a human single-domain antibody that spe- cifically recognized amyloid-beta 42 oligomers was identified. Western blot and enzyme-linked im- munosorbent assay demonstrated this antibody bound specifically to human amyloid-beta 42 tetramer and nonamer, but not the monomer or high molecular weight oligomers. This study suc- cessfully constructed a human phage display library and screened a single-domain antibody that specifically recognized amyloid-beta 42 oligomers.

Maternal-derived antibodies hinder the antibody response to H9N2 AIV inactivated vaccine in the field

The H9N2 subtype avian influenza virus(AIV)inactivated vaccine has been used extensively in poultry farms,but it often fails to stimulate a sufficiently high immune response in poultry in the field,although it works well in laboratory experiments;hence,the virus still causes economic damage every year and poses a potential threat to public health.Based on surveillance data collected in the field,we found that broilers with high levels of maternal-derived antibodies(MDAs)against H9N2 virus did not produce high levels of antibodies after vaccination with a commercial H9N2 inactivated vaccine.In contrast,specific pathogen-free(SPF)chickens without MDAs responded efficiently to that vaccination.When MDAs were mimicked by administering passively transferred antibodies(PTAs)into SPF chickens in the laboratory,similar results were observed:H9N2-specific PTAs inhibited humoral immunity against the H9N2 inactivated vaccine,suggesting that H9N2-specific MDAs might hinder the generation of antibodies when H9N2 inactivated vaccine was used.After challenge with homologous H9N2 virus,the virus was detected in oropharyngeal swabs of the vaccinated and unvaccinated chickens with PTAs but not in the vaccinated chickens without PTAs,indicating that H9N2-specific MDAs were indeed one of the reasons for H9N2 inactivated vaccine failure in the field.When different titers of PTAs were used to mimic MDAs in SPF chickens,high(HI=12 log2)and medium(HI=log 9 log2)titers of PTAs reduced the generation of H9N2-specific antibodies after the first vaccination,but a booster dose would induce a high and faster humoral immune response even of PTA interference.This study strongly suggested that high or medium titers of MDAs might explain H9N2 inactivated vaccine failure in the field.