Multiplexed immunohistochemistry/fluorescence(mIHC/IF)in combination with multispectral unmixing is a novel multitarget histopathological staining and imaging technique.By simultaneously revealing expression level and...Multiplexed immunohistochemistry/fluorescence(mIHC/IF)in combination with multispectral unmixing is a novel multitarget histopathological staining and imaging technique.By simultaneously revealing expression level and spatial information for up to eight biomarkers in situ,in addition to a nuclear stain within a single formalin-fixed paraffin-embedded(FFPE)tissue section,this technology can analyze the phenotype,abundance,morphology and intercellular relationship of cells while providing statistically significant results.In recent years,technical improvements have brought new insight into mIHC/IF and multispectral imaging approaches,which have been successfully applied in the field of cancer immunotherapy,specifically in regard to tumor microenvironment research,immunotherapy drug discovery,and prognostic and metastatic risk evaluation.This review highlights the principle,workflow,advantages and disadvantages of the technology,and discusses the latest applications of mIHC/IF-based imaging technology in the field of TME-related research and immunotherapy drug discovery.展开更多
A total of 66 samples （from 27 cases with neuromyelitis optica, 26 cases with multiple sclerosis, aa 13 cases with optic neuritis） were tested for aquaporin-4 antibody by a cell-based immunofluorescence assay and an...A total of 66 samples （from 27 cases with neuromyelitis optica, 26 cases with multiple sclerosis, aa 13 cases with optic neuritis） were tested for aquaporin-4 antibody by a cell-based immunofluorescence assay and an enzyme-linked immunosorbent assay. The sensitivities and specificities of the two assays were similar. We further analyzed an additional 68 patients and 93 healthy controls using the enzyme-linked immunosorbent assay. A Kappa test showed good consistency between the two methods in terms of detection of anti-aquaporin-4 antibody in the se of neuromyelitis optica patients. No significant correlations were identified with onset age or disea duration, suggesting that aquaporin-4 antibody is a good marker for neuromyelitis optica. The enzyme-linked immunosorbent assay can be used for quantifying aquaporin-4 antibody concentrations and may be useful to dynamically monitor changes in the levels of aquaporin-4 antibody during disease duration.展开更多
AIM： To evaluate the novel anti-endomysium （anti-EMA） detection based on ELISA. METHODS： Anti-EMA IgA was measured by a novel ELISA in 196 patients with gastrointestinal symptoms and suspected mal-absorption. Data...AIM： To evaluate the novel anti-endomysium （anti-EMA） detection based on ELISA. METHODS： Anti-EMA IgA was measured by a novel ELISA in 196 patients with gastrointestinal symptoms and suspected mal-absorption. Data were compared with those obtained by the conventional IF test. RESULTS： A good concordance of 98% was found between these two assays. In sera of 161 patients （82%） both assays tested negative whereas in sera of 31 patients （16%） both assays tested positive for the presence of anti-EMA antibodies. Discrepancies between EMA- ELISA and EMA-immunofluorescence （IF） were found in only 4 patients （2%）.CONCLUSION： This ELISA can replace IF for the detection of anti-EMA antibodies and provide clinicians with an excellent tool to screen for celiac disease in patients with gastrointestinal complaints.展开更多
AIMTo describe the technique of immunofluorescence on paraffin embedded tissue sections and discuss the po-tential pitfalls with an in depth review of literature.METHODSImmunofluorescence is integral to diagnostic ren...AIMTo describe the technique of immunofluorescence on paraffin embedded tissue sections and discuss the po-tential pitfalls with an in depth review of literature.METHODSImmunofluorescence is integral to diagnostic renal pa-thology. Immunofluorescence on paraffin embedded renal biopsies （IF-P） after enzyme treatment has been described in literature, however has not found widespread use in renal pathology laboratories. In our laboratory proteinase K digestion of paraffn embedded renal biopsy material was standardized and applied prospectively in cases where immunofuorescence on fresh frozen tissue was non contributory or not possible. Diagnostic utility was assessed and in a cohort of cases comparison of intensity of staining with routine immunofuorescence was performed.RESULTSOver the 5-year study period, of the 3141 renal biopsies received IF-P was performed on 246 cases （7.7%） and was interpretable with optimal digestion in 214 cases （6.8%）. It was of diagnostic utility in the majority of cases, which predominantly included glomerular disease. Non-diagnostic IF-P was found in membranous nephropathy （2 of 11 cases）, membranoproliferative glomerulonephritis （2 of 32 cases）, lupus nephritis （1 of 25 cases）, post infectious glomerulonephritis （1 of 11 cases） and chronic glomerulonephritis （3 of 8 cases）. Comparing cases with both routine IF and IF-P, 35 of 37 showed either equal intensity or a minor difference in intensity of staining（1＋） for the diagnostic immunoglobulin/complement. Technically assessment of immunofluorescence on the paraffin embedded tissue was found to be easier with clearly observed morphology, however a false positive staining pattern was observed in under-digested tissue. CONCLUSIONAs a “salvage” technique, immunofuorescence on paraffn embedded renal biopsies is of great diagnostic utility, however not without pitfalls.展开更多
Objective To compare the effectiveness of immunofluorescence and polymerase chain reaction （PCR） in detecring human papilloma virus （HPV） in condyloma acuminata （CA）. Methods HPVs in CA tissues from 60 patients...Objective To compare the effectiveness of immunofluorescence and polymerase chain reaction （PCR） in detecring human papilloma virus （HPV） in condyloma acuminata （CA）. Methods HPVs in CA tissues from 60 patients were detected by immunofluorescence and PCR, respectively. Different subtypes of HPVs were also identified with restriction fragment length polymorphism （RFLP） . Resuits The positive detective rates of immunofluorescence and PCR were 56. 67 % （34/60） and 96. 67 % （58/ 60）, respectively （ P 〈 0. 01 ）. RFLP results showed HPV6 and HPV11 were the main subtypes in the detected virus, which accounted for 98.28%. Conclusion The sensibility of PCR is superior to that of immunofluorescence.展开更多
Objective: The aim was to detect the expression of PR and CD146 in paraf-fin-embedded tissue sections of endometrioid adenocarcinoma by using QDs double-labeling immunofluorescence, and evaluate the applied value of Q...Objective: The aim was to detect the expression of PR and CD146 in paraf-fin-embedded tissue sections of endometrioid adenocarcinoma by using QDs double-labeling immunofluorescence, and evaluate the applied value of QDs double-labeling immunofluorescence in endometrioid adenocarcinoma. Methods: To detect the expression of PR and CD146 on 140 cases of paraffin-embedded tissue sections of endometrioid adenocarcinoma by using QDS double-labeling immunofluorescence. Results: The co-expression of PR and CD146 in the endometrioid adenocarcinoma can be detected by QDs double-labeling immunofluorescence, and there was no correlation between them (P > 0.05). Conclusion: QDs double-labeling immunofluorescence can detect the localization and co-expression of PR and CD146 in the endometrioid adenocarcinoma.展开更多
BACKGROUND Serologic cross-reactivity between hantaviruses often complicates the interpretation of the results.AIM To analyze the diagnostic value of indirect immunofluorescence assay(IFA)and western blot(WB)in the di...BACKGROUND Serologic cross-reactivity between hantaviruses often complicates the interpretation of the results.AIM To analyze the diagnostic value of indirect immunofluorescence assay(IFA)and western blot(WB)in the diagnosis of hantavirus infections.METHODS One hundred eighty-eight serum samples from Puumala(PUUV)and Dobrava(DOBV)orthohantavirus infected patients were analyzed.Serology was performed using commercial tests(Euroimmun,Lübeck,Germany).RESULTS Using IFA,49.5%of acute-phase samples showed a monotypic response to PUUV,while 50.5% cross-reacted with other hantaviruses.The overall cross-reactivity was higher for immunoglobulin G(IgG)(50.0%)than for immunoglobulin M(IgM)(25.5%).PUUV IgM/IgG antibodies showed low/moderate reactivity with orthohantaviruses Hantaan(12.3%/31.5%),Seoul(7.5%/17.8%),DOBV(5.4%/28.1%),and Saaremaa(4.8%/15.7%).Both DOBV IgM and IgG antibodies were broadly reactive with Hantaan(76.2%/95.2%),Saaremaa(80.9%/83.3%),and Seoul(78.6%/85.7%)and moderate with PUUV(28.5%/38.1%).Using a WB,serotyping was successful in most cross-reactive samples(89.5%).CONCLUSION The presented results indicate that WB is more specific than IFA in the diagnosis of hantavirus infections,confirming serotype in most IFA cross-reactive samples.展开更多
ASY （asynaptic） is a synaptonemal complex （SC） related protein in plant mei- osis. Some ASYs in plants have been cloned and functional determined. ASY in Brassica napus has not been sequenced and functional certi...ASY （asynaptic） is a synaptonemal complex （SC） related protein in plant mei- osis. Some ASYs in plants have been cloned and functional determined. ASY in Brassica napus has not been sequenced and functional certificated. Here, we first cloned ASY gene in Brassica napus （BnASY） and inserted it into vector pET-32a for expression fusion protein BnASY-6His in Escherichia coli BL21 （DE3）. Purified fusion protein was used to produce polyclonal antibody. Specificity and application of polyclonal antibody was examined by Western blot and immunofluorescence assay. Results showed that BnASY-6His recombi-nant protein and anti-BnASY antibody were successfully obtained. Polyclonal anti-BnASY antibody had high specificity. Results of immunofluorescence showed that ASY signals appeared at leptotene stage, faded away gradually at pachytene stage and almost disap-peared at diplonema stage. In this study, highly specific polyclonal antibody was success-fully acquired and used for immunofluorescence assay in plant cells. It will contribute to functional research of ASY in B. napus.We thank Zhigang Li for providing pET-32a vector and Escherichia coli BL21 pLysS strains.展开更多
Conventional immunohistochemistry(IHC)is a widely used diagnostic technique in tissue pathology.However,this technique is associated with a number of limitations,including high inter-observer variability and the capac...Conventional immunohistochemistry(IHC)is a widely used diagnostic technique in tissue pathology.However,this technique is associated with a number of limitations,including high inter-observer variability and the capacity to label only one marker per tissue section.This review details various highly multiplexed techniques that have emerged to circumvent these constraints,allowing simultaneous detection of multiple markers on a single tissue section and the comprehensive study of cell composition,cellular functional and cell-cell interactions.Among these techniques,multiplex Immunohistochemistry/Immunofluorescence(mIHC/IF)has emerged to be particularly promising.mIHC/IF provides high-throughput multiplex staining and standardized quantitative analysis for highly reproducible,efficient and cost-effective tissue studies.This technique has immediate potential for translational research and clinical practice,particularly in the era of cancer immunotherapy.展开更多
Immunofluorescence has been widely used to localize microbes or specific molecules in insect tissues or cells.However,significant autofluorescence is frequently observed in tissues which can interfere with the fluores...Immunofluorescence has been widely used to localize microbes or specific molecules in insect tissues or cells.However,significant autofluorescence is frequently observed in tissues which can interfere with the fluorescent identification of target antigens,leading to inaccurate or even false positive fluorescent labeling.The alimentary canal of the potato psyllid,Bactericera cockerelliŠulc,exhibits intense autofluorescence,hindering the application of immunolocalization for the detection and localization of the economically important pathogen transmitted by this insect,“Candidatus Liberibacter solanacearum”(Lso).In the present study,we tested the use of irradiation,hydrogen peroxide(H2O2)and Sudan black B(SBB)treatments to reduce the autofluorescence in the B.cockerelli alimentary canal tissues.Furthermore,we assessed the compatibility of the above‐mentioned treatments with Lso immunolocalization and actin staining using phalloidin.Our results showed that the autofluorescence in the alimentary canal was reduced by irradiation,H2O2,or SBB treatments.The compatibility assays indicated that irradiation and H2O2 treatment both greatly reduced the fluorescent signal associated with Lso and actin.However,the SBB incubation preserved those target signals,while efficiently eliminating autofluorescence in the psyllid alimentary canal.Therefore,herein we propose a robust method for reducing the autofluorescence in the B.cockerelli alimentary canal with SBB treatment,which may improve the use of immunofluorescence labeling in this organism.This method may also have a wide range of uses by reducing the autofluorescence in other arthropod species.展开更多
Xyloglucans in the non-lignified primary cell walls of different species of monocotyledons have diverse struc- tures, with widely varying proportions of oligosaccharide units that contain fucosylated side chains （F s...Xyloglucans in the non-lignified primary cell walls of different species of monocotyledons have diverse struc- tures, with widely varying proportions of oligosaccharide units that contain fucosylated side chains （F side chains）. To determine whether fucosylated xyloglucans occur in all non-lignified walls in a range of monocotyledon species, we used immunofluorescence microscopy with the monoclonal antibody CCRC-M1. The epitope of this antibody, α-L-FUCp-（1 →2）- β-D-Galp, occurs in F side chains. In most non-commelinid monocotyledons, the epitope was found in all non-lignified walls. A similar distribution was found in the palm Phoenix canariensis, which is a member of the basal commelinid order Arecales. However, in the other commelinid orders Zingiberales, Commelinales, and Poales, the occurrence of the epitope was restricted, sometimes occurring in only the phloem walls, but often also in walls of other cell types including stomatal guard and subsidiary cells and raphide idioblasts. No epitope was found in the walls of the commelinids Tradescantia virginiana （Commelinaceae, Commelinales） and Zea mays （Poaceae, Poales）, but it occurred in the phloem walls of two other Poaceae species, Lolium multiflorum and L. perenne. The distribution of the epitope is discussed in relation to xyloglucan structures in the different taxa. However, the functional significance of the restricted distributions is unknown.展开更多
Erratum to:J Huazhong Univ Sci Technol[Med Sci]36(4):548–553,2016 https://doi.org/10.1007/s11596-016-1623-6 In the originally published article(https://doi.org/10.1007/s11596-016-1623-6),the immunofluorescence images...Erratum to:J Huazhong Univ Sci Technol[Med Sci]36(4):548–553,2016 https://doi.org/10.1007/s11596-016-1623-6 In the originally published article(https://doi.org/10.1007/s11596-016-1623-6),the immunofluorescence images in shRNA group in Fig.3 were accidentally used rather than the final,formal experiments.To retain consistency,the entire Fig.3 is replaced here with original images of the experiments.The authors declare that this correction will not affect the conclusion of the study.展开更多
With the deepening of epigenetic research,studies have shown that N6-methyladenosine(m^(6)A)is closely related to the development of rheumatoid arthritis(RA),but the mechanism is still unclear.In the study,we collecte...With the deepening of epigenetic research,studies have shown that N6-methyladenosine(m^(6)A)is closely related to the development of rheumatoid arthritis(RA),but the mechanism is still unclear.In the study,we collected synovial tissues from normal controls and patients with osteoarthritis(OA)or RA.The levels of m^(6)A and inflammation were analyzed by immunofluorescence staining and western blotting.The roles of IGF2BP3 in cell proliferation and inflammatory activation were explored using transfection and RNA immunoprecipitation assays.IGF2BP3^(−/−)mice were generated and used to establish an arthritis mouse model by transferring serum from adult arthritis K/BxN mice.We found m^(6)A levels were markedly increased in RA patients and mouse models,and the expression of IGF2BP3 was upregulated in individuals with RA and related to the levels of inflammatory markers.IGF2BP3 played an important part in RA-fibroblast-like synoviocytes(FLS)by promoting cell proliferation,migration,invasion,inflammatory cytokine release and inhibiting autophagy.In addition,IGF2BP3 inhibited autophagy to reduce ROS production,thereby decreasing the inflammatory activation of macrophages.More importantly,RASGRF1-mediated mTORC1 activation played a crucial role in the ability of IGF2BP3 to promote cell proliferation and inflammatory activation.In an arthritis model of IGF2BP3^(−/−)mice,IGF2BP3 knockout inhibited RA-FLS proliferation and inflammatory infiltration,and further ameliorated RA joint injury.Our study revealed an important role for IGF2BP3 in RA progression.The targeted inhibition of IGF2BP3 reduced cell proliferation and inflammatory activation and limited RA development,providing a potential strategy for RA therapy.展开更多
BACKGROUND Patients with tumors often develop multiple autoantibodies against tumorassociated antigens.Among these,antinuclear antibodies(ANAs)constitute a clinically important group distributed across the nucleus,cyt...BACKGROUND Patients with tumors often develop multiple autoantibodies against tumorassociated antigens.Among these,antinuclear antibodies(ANAs)constitute a clinically important group distributed across the nucleus,cytoplasm,and cytoskeleton.Emerging evidence suggests that ANAs are closely associated with the development and progression of various malignancies,including colorectal cancer(CRC).AIM To detect ANA fluorescence patterns in CRC using indirect immunofluorescence(IIF)and investigate their correlation with the disease.METHODS We collected serum samples from patients and healthy controls visiting The Affiliated Panyu Central Hospital of Guangzhou Medical University between May 2023 and March 2024 for analysis.The study included 38 patients with newly diagnosed CRC,43 patients with colorectal polyps(CRP),and 29 healthy controls.Serum ANA expression was assessed by IIF,and fluorescence patterns were recorded for each group.Differences in ANA titers were compared among each group to analyze the differences in serum ANA-positive expression,which were further analyzed to explore the correlation between ANA expression and CRC screening.RESULTS ANA positivity rates were 50.00%in the CRC group,46.51%in the colorectal polyp group,and 6.90%in the healthy control group,with significantly higher rates in the two patient groups compared to the control group(P<0.05).In the CRC group,the most common fluorescence patterns were nuclear speckled(15.79%)and cytoplasmic speckled(15.79%),with titers predominantly low(1:100,28.95%).In the colorectal polyp group,nuclear speckled(18.60%)and nuclear homogeneous(11.63%)were the most frequent,with titers also predominantly low(1:100,37.21%).The distribution of intermediate titers differed significantly among groups(P<0.05).CONCLUSION ANAs are associated with both CRP and CRC and may be useful in early CRC screening.Medium-to-high ANA titers,in particular,should prompt further evaluation for possible CRC correlation.Multiple ANA fluorescence patterns can be detected across all groups,with patients with CRP and CRC showing greater pattern diversity than healthy controls.展开更多
[Objective] The aim of the present study was to develop a direct immunofluorescence method for the diagnosis of canine distemper （CD） with FITC-conjugated monoclonal antibodies （FITC-McAb）.[ Metbod] The McAb again...[Objective] The aim of the present study was to develop a direct immunofluorescence method for the diagnosis of canine distemper （CD） with FITC-conjugated monoclonal antibodies （FITC-McAb）.[ Metbod] The McAb against CDV, designated as CE3, was purified with protein G and labeled with FITC through agitation method. After purification and identification, the optimal working concentration of FITC-labeled CE3 was determined. Then 61 clinical samples of suspected canine distemper were detected by direct immunofluorescence assay. [ Result] The absorption test, blocking test and specificity test showed that the labeled antibody had high specificity and sensitivity, but didn＇t have cross reaction with canine parvovirus （CPV）, canine parainfluenza virus （CPIV）, canine adenovirus （CAV） and rabies virus （RV）. The optimal working concentration was 1：80. The positive rate of clinical suspected samples was 48%. [ Conclusion] The direct immunofluorescence assay developed in this study was rapid, specific and convenient, and had great significance for the early diagnosis of canine distemper.展开更多
Background: Diarrheal diseases have globally decreased over the past few decades, yet they remain one of the top three causes of mortality in children under five years, especially in sub-Saharan Africa and Nigeria. Se...Background: Diarrheal diseases have globally decreased over the past few decades, yet they remain one of the top three causes of mortality in children under five years, especially in sub-Saharan Africa and Nigeria. Seasonal peaks of diarrheal episodes continue to contribute significantly to childhood mortality in these regions. One of the notable causes of diarrhea in children is parasitic infections, particularly Cryptosporidium, which poses a serious health risk. In Nigeria, the burden of Cryptosporidium diarrhea is under-researched, making it imperative to investigate its prevalence and associated risk factors. Study Objectives: The study aims to determine the prevalence and risk factors associated with Cryptosporidium diarrhea among children aged five years and below in selected health institutions in Abakaliki, South-East Nigeria. Methodology: This cross-sectional study was conducted from January to May 2017, recruiting 200 children under five years with diarrhea from health institutions in Abakaliki. Fecal specimens were analyzed for Cryptosporidium oocysts using light microscopy with modified Ziehl-Neelsen staining and immunofluorescent antibody test (IFAT). Deoxyribonucleic acid (DNA) was extracted from positive samples using QIAmp® DNA stool kit, followed by Polymerase Chain Reaction (PCR) and molecular genotyping. Results: Cryptosporidium was detected in 0.5% (1/200) of children via light microscopy and 6.5% (13/200) via IFAT. All positive samples were confirmed as Cryptosporidium hominis by PCR. The prevalence of infection was significantly higher in children from institutionalized homes (50.0%) compared to monogamous homes (6.2%) (p Conclusion: Cryptosporidium hominis is a notable cause of diarrhea among children in Abakaliki, primarily transmitted through human-to-human contact. The study underscores the need for targeted interventions in childcare institutions to prevent outbreaks. Health authorities should promote breastfeeding and enhance education on hygiene practices in vulnerable populations.展开更多
Objective: To study the relationship between Chlamydia pneumoniae (C. pneumoniae) infection and asthma exacerbation. Methods: A prospective study of C. pneumoniae infection was conducted in 75 patients with asthma and...Objective: To study the relationship between Chlamydia pneumoniae (C. pneumoniae) infection and asthma exacerbation. Methods: A prospective study of C. pneumoniae infection was conducted in 75 patients with asthma and 63 patients with respiratory tract infection, and 100 blood donors served as controls. The presence of infection was convinced by the polymerase chain reaction and direct immunofluorescence assay for C. pneumoniae DNA from throat swab specimens and micro-immunofluorescence testing for C. pneu-moniae-specific IgG, IgM and IgA antibodies. Results: Prevalence of specific IgG in asthma patients (81. 3%) was higher than that of the blood donors (68. 0%, P<0. 05) and was not significantly different from respiratory tract infection patients (68. 0%, P>0. 05). The acute C. pneumoniae infection rate of symptomatic asthma patients (59. 4%) was markedly higher than that of respiratory tract infection patients (34. 9% , P<0. 05). The average titer of C. pneumoniae IgG instead of IgA in asthma patients (48. 38±6. 94) was significantly higher than respiratory tract infection patients (24. 70±8. 77, P<0. 05). Other pathogens were identified in 12 of 21 (57. 1%) asthma patients with C. pneumoniae. The symptoms of 7 asthma patients with C. pneumoniae infection were improved through antibiotic treatment. Conclusion: The findings suggest a possible role of C. pneumoniae infection in asthma.展开更多
One of the prominent cell cycle related modifications of histone proteins, whose function is correlated with chromosome condensation, is the phosphorylation of histone H3. Wheat (Triticum aestivum L.) mitotic and meio...One of the prominent cell cycle related modifications of histone proteins, whose function is correlated with chromosome condensation, is the phosphorylation of histone H3. Wheat (Triticum aestivum L.) mitotic and meiotic cells were analyzed with indirect immunoflurorescence labeling with an antibody recognizing histone H3 phosphorylated at Serine 10 to study the localization of phosphorylated histone H3 at mitosis and meiosis. Our results showed that, during mitotic division, the phosphoryiation of H3 started from early prophase and vanished at telophase, remaining mainly in the pericentromeric regions at metaphase and anaphase. During meiotic division, phosphorylation of H3 initiated at the transition from leptotene to zygotene and remained uniform, along the chromosomes from prophase I until telophase whereas it showed slightly stronger in the pericentromeric regions than along the chromosome arms from metaphase II until Lelophase II The different patterns of H3 phophorylation at mitosis and meiosis in wheat suggested that this evolutionarily conserved post-translational chromatin modification might be involved in more roles besides chromosome condensation.展开更多
Five monoclonal antibodies（Mabs） to nuclear protein of avain influenza virus（AIV） were developed by syncretizing SP 2/0 and the spleen cells from BALB of mice immuized with H9 subtype AIV. Specificity of these Mab...Five monoclonal antibodies（Mabs） to nuclear protein of avain influenza virus（AIV） were developed by syncretizing SP 2/0 and the spleen cells from BALB of mice immuized with H9 subtype AIV. Specificity of these Mabs were identified by immunofluorescent assay（IFA） and enzyme linked immunosorbent assay （ELISA）. These five Mabs which were named as AIV-NP-2C3, AIV-NP-6A5, AIV-NP-3 H9, AIV-NP-7B4, AIV-NP-2H4 could react with all viruses of AIV-H9 strains in tests. The result of Western blotting showed that only the 60 ku protein antigen of AIV-H9 could be recognized by the Mabs but never recognized by New castle disease virus, REV and infectious bursa disease virus. The result of preliminary application showed that avian influenza viruses could be deetected bv Mabs in IFA and ELISA. All these Mabs will probably play important roles in preventing and monitoring avian influenza viruses.展开更多
AIM: To investigate whether KAI1, as a metastasis suppressor gene, is associated with invasive and metastatic ability of pancreatic cancer cells.METHODS: KAI1 gene was transfected into pancreatic cancer cell line MiaP...AIM: To investigate whether KAI1, as a metastasis suppressor gene, is associated with invasive and metastatic ability of pancreatic cancer cells.METHODS: KAI1 gene was transfected into pancreatic cancer cell line MiaPaCa Ⅱ by liposomes selected with G418. Expression of transfected cells was measured by Western blotting, immunofluorescence and immunocytochemistry. Tumor cell invasion and metastatic ability were detected through gelatinase activity and reconstituted basement membrane (Matrigel) assay. pCMV-KAI1 was directly injected into the heterotopic human pancreatic adenocarcinoma successfully established in the groin of BALB/C nude mice, by subcutaneous injection of MiaPaCa Ⅱ pancreatic cancer cells. The statistical analysis between groups was determined by Student's two tailed t test.RESULTS: By Western blotting, MiaPaCa Ⅱ cells transfected by KAI1 gene indicated KAI1 expression at approximately 29.1 kDa. Cytoplasm staining was positive and uniformly spread in transfected cancer cells, using immunohistochemistry and immunofluorescence. The most obvious difference was present after 30 h (MiaPaca Ⅱ 43.6 ± 9.42, pCMV-MiaPaca Ⅱ 44.8 ± 8.56, pCMV-KAI1-MiaPaca Ⅱ 22.0 ± 4.69, P < 0.05). Gelatinolysis revealed a wider and clearer band of gelatinolytic activity in non-transfected than in transfected cells (MiaPaCa Ⅱ cells 30.8 ± 0.57, transfected cells 28.1 ± 0.65, P < 0.05). In vivo tumor growth rates of KAI1 transfectants with KAI1-Lipofectamine 1.22 ± 0.31 in A group were lower than control 4.61 ± 1.98 and pCMV-KAI 11.67 ± 0.81. Analyses of metastases with and without KAI1 transfection in mice were different in liver and lung between controls 1.62 ± 0.39, 0.45 ± 0.09, pCMV-KAI 1.01 ± 0.27, 0.33 ± 0.09 and KAI1-Lipofectamine 0.99 ± 0.21, 0.30 ± 0.09 respectively (P < 0.05).CONCLUSION: High expression of KAI1 gene was found in transfected MiaPaCa Ⅱ human pancreatic cancer cells with lower metastatic ability. KAI1 gene plays an important role in inhibiting metastasis of pancreatic cancer after direct injection into pancreatic adenocarcinoma. These results show that the suppressed invasion and motor function of pancreatic cancer cells may be a key reason why the KAI1 gene controls pancreatic cancer cell metastasis.展开更多
基金supported by State Key Laboratory of Natural and Biomimetic Drugs,Peking University。
文摘Multiplexed immunohistochemistry/fluorescence(mIHC/IF)in combination with multispectral unmixing is a novel multitarget histopathological staining and imaging technique.By simultaneously revealing expression level and spatial information for up to eight biomarkers in situ,in addition to a nuclear stain within a single formalin-fixed paraffin-embedded(FFPE)tissue section,this technology can analyze the phenotype,abundance,morphology and intercellular relationship of cells while providing statistically significant results.In recent years,technical improvements have brought new insight into mIHC/IF and multispectral imaging approaches,which have been successfully applied in the field of cancer immunotherapy,specifically in regard to tumor microenvironment research,immunotherapy drug discovery,and prognostic and metastatic risk evaluation.This review highlights the principle,workflow,advantages and disadvantages of the technology,and discusses the latest applications of mIHC/IF-based imaging technology in the field of TME-related research and immunotherapy drug discovery.
基金the grants from the Ministry of Sciences and Technology of China, No. 2006AA02A408, 2008ZX09312-014
文摘A total of 66 samples （from 27 cases with neuromyelitis optica, 26 cases with multiple sclerosis, aa 13 cases with optic neuritis） were tested for aquaporin-4 antibody by a cell-based immunofluorescence assay and an enzyme-linked immunosorbent assay. The sensitivities and specificities of the two assays were similar. We further analyzed an additional 68 patients and 93 healthy controls using the enzyme-linked immunosorbent assay. A Kappa test showed good consistency between the two methods in terms of detection of anti-aquaporin-4 antibody in the se of neuromyelitis optica patients. No significant correlations were identified with onset age or disea duration, suggesting that aquaporin-4 antibody is a good marker for neuromyelitis optica. The enzyme-linked immunosorbent assay can be used for quantifying aquaporin-4 antibody concentrations and may be useful to dynamically monitor changes in the levels of aquaporin-4 antibody during disease duration.
文摘AIM： To evaluate the novel anti-endomysium （anti-EMA） detection based on ELISA. METHODS： Anti-EMA IgA was measured by a novel ELISA in 196 patients with gastrointestinal symptoms and suspected mal-absorption. Data were compared with those obtained by the conventional IF test. RESULTS： A good concordance of 98% was found between these two assays. In sera of 161 patients （82%） both assays tested negative whereas in sera of 31 patients （16%） both assays tested positive for the presence of anti-EMA antibodies. Discrepancies between EMA- ELISA and EMA-immunofluorescence （IF） were found in only 4 patients （2%）.CONCLUSION： This ELISA can replace IF for the detection of anti-EMA antibodies and provide clinicians with an excellent tool to screen for celiac disease in patients with gastrointestinal complaints.
文摘AIMTo describe the technique of immunofluorescence on paraffin embedded tissue sections and discuss the po-tential pitfalls with an in depth review of literature.METHODSImmunofluorescence is integral to diagnostic renal pa-thology. Immunofluorescence on paraffin embedded renal biopsies （IF-P） after enzyme treatment has been described in literature, however has not found widespread use in renal pathology laboratories. In our laboratory proteinase K digestion of paraffn embedded renal biopsy material was standardized and applied prospectively in cases where immunofuorescence on fresh frozen tissue was non contributory or not possible. Diagnostic utility was assessed and in a cohort of cases comparison of intensity of staining with routine immunofuorescence was performed.RESULTSOver the 5-year study period, of the 3141 renal biopsies received IF-P was performed on 246 cases （7.7%） and was interpretable with optimal digestion in 214 cases （6.8%）. It was of diagnostic utility in the majority of cases, which predominantly included glomerular disease. Non-diagnostic IF-P was found in membranous nephropathy （2 of 11 cases）, membranoproliferative glomerulonephritis （2 of 32 cases）, lupus nephritis （1 of 25 cases）, post infectious glomerulonephritis （1 of 11 cases） and chronic glomerulonephritis （3 of 8 cases）. Comparing cases with both routine IF and IF-P, 35 of 37 showed either equal intensity or a minor difference in intensity of staining（1＋） for the diagnostic immunoglobulin/complement. Technically assessment of immunofluorescence on the paraffin embedded tissue was found to be easier with clearly observed morphology, however a false positive staining pattern was observed in under-digested tissue. CONCLUSIONAs a “salvage” technique, immunofuorescence on paraffn embedded renal biopsies is of great diagnostic utility, however not without pitfalls.
文摘Objective To compare the effectiveness of immunofluorescence and polymerase chain reaction （PCR） in detecring human papilloma virus （HPV） in condyloma acuminata （CA）. Methods HPVs in CA tissues from 60 patients were detected by immunofluorescence and PCR, respectively. Different subtypes of HPVs were also identified with restriction fragment length polymorphism （RFLP） . Resuits The positive detective rates of immunofluorescence and PCR were 56. 67 % （34/60） and 96. 67 % （58/ 60）, respectively （ P 〈 0. 01 ）. RFLP results showed HPV6 and HPV11 were the main subtypes in the detected virus, which accounted for 98.28%. Conclusion The sensibility of PCR is superior to that of immunofluorescence.
文摘Objective: The aim was to detect the expression of PR and CD146 in paraf-fin-embedded tissue sections of endometrioid adenocarcinoma by using QDs double-labeling immunofluorescence, and evaluate the applied value of QDs double-labeling immunofluorescence in endometrioid adenocarcinoma. Methods: To detect the expression of PR and CD146 on 140 cases of paraffin-embedded tissue sections of endometrioid adenocarcinoma by using QDS double-labeling immunofluorescence. Results: The co-expression of PR and CD146 in the endometrioid adenocarcinoma can be detected by QDs double-labeling immunofluorescence, and there was no correlation between them (P > 0.05). Conclusion: QDs double-labeling immunofluorescence can detect the localization and co-expression of PR and CD146 in the endometrioid adenocarcinoma.
文摘BACKGROUND Serologic cross-reactivity between hantaviruses often complicates the interpretation of the results.AIM To analyze the diagnostic value of indirect immunofluorescence assay(IFA)and western blot(WB)in the diagnosis of hantavirus infections.METHODS One hundred eighty-eight serum samples from Puumala(PUUV)and Dobrava(DOBV)orthohantavirus infected patients were analyzed.Serology was performed using commercial tests(Euroimmun,Lübeck,Germany).RESULTS Using IFA,49.5%of acute-phase samples showed a monotypic response to PUUV,while 50.5% cross-reacted with other hantaviruses.The overall cross-reactivity was higher for immunoglobulin G(IgG)(50.0%)than for immunoglobulin M(IgM)(25.5%).PUUV IgM/IgG antibodies showed low/moderate reactivity with orthohantaviruses Hantaan(12.3%/31.5%),Seoul(7.5%/17.8%),DOBV(5.4%/28.1%),and Saaremaa(4.8%/15.7%).Both DOBV IgM and IgG antibodies were broadly reactive with Hantaan(76.2%/95.2%),Saaremaa(80.9%/83.3%),and Seoul(78.6%/85.7%)and moderate with PUUV(28.5%/38.1%).Using a WB,serotyping was successful in most cross-reactive samples(89.5%).CONCLUSION The presented results indicate that WB is more specific than IFA in the diagnosis of hantavirus infections,confirming serotype in most IFA cross-reactive samples.
基金This work was supported by National Natural Science Foundation of China （31671733,31400243 and 31201152）, Natural Science Foundation of Hubei Province （2013CFB423 and 2014CFB320）, Major Research Project of CAAS Science, Technology Innovation Program and the Cen-tral Public-interest Scientific Institution Basal Research Fund.
文摘ASY （asynaptic） is a synaptonemal complex （SC） related protein in plant mei- osis. Some ASYs in plants have been cloned and functional determined. ASY in Brassica napus has not been sequenced and functional certificated. Here, we first cloned ASY gene in Brassica napus （BnASY） and inserted it into vector pET-32a for expression fusion protein BnASY-6His in Escherichia coli BL21 （DE3）. Purified fusion protein was used to produce polyclonal antibody. Specificity and application of polyclonal antibody was examined by Western blot and immunofluorescence assay. Results showed that BnASY-6His recombi-nant protein and anti-BnASY antibody were successfully obtained. Polyclonal anti-BnASY antibody had high specificity. Results of immunofluorescence showed that ASY signals appeared at leptotene stage, faded away gradually at pachytene stage and almost disap-peared at diplonema stage. In this study, highly specific polyclonal antibody was success-fully acquired and used for immunofluorescence assay in plant cells. It will contribute to functional research of ASY in B. napus.We thank Zhigang Li for providing pET-32a vector and Escherichia coli BL21 pLysS strains.
文摘Conventional immunohistochemistry(IHC)is a widely used diagnostic technique in tissue pathology.However,this technique is associated with a number of limitations,including high inter-observer variability and the capacity to label only one marker per tissue section.This review details various highly multiplexed techniques that have emerged to circumvent these constraints,allowing simultaneous detection of multiple markers on a single tissue section and the comprehensive study of cell composition,cellular functional and cell-cell interactions.Among these techniques,multiplex Immunohistochemistry/Immunofluorescence(mIHC/IF)has emerged to be particularly promising.mIHC/IF provides high-throughput multiplex staining and standardized quantitative analysis for highly reproducible,efficient and cost-effective tissue studies.This technique has immediate potential for translational research and clinical practice,particularly in the era of cancer immunotherapy.
基金This work was supported by Texas A&M University and Texas A&M AgriLife Research(Controlling Exotic and Invasive Insect-Transmitted Pathogens)and Hatch project TEX0-1-9381 Accession Number 1015773Xiaotian Tang received the Herb Dean'40 Endowed Scholarship from the Department of Entomology at Texas A&M University.
文摘Immunofluorescence has been widely used to localize microbes or specific molecules in insect tissues or cells.However,significant autofluorescence is frequently observed in tissues which can interfere with the fluorescent identification of target antigens,leading to inaccurate or even false positive fluorescent labeling.The alimentary canal of the potato psyllid,Bactericera cockerelliŠulc,exhibits intense autofluorescence,hindering the application of immunolocalization for the detection and localization of the economically important pathogen transmitted by this insect,“Candidatus Liberibacter solanacearum”(Lso).In the present study,we tested the use of irradiation,hydrogen peroxide(H2O2)and Sudan black B(SBB)treatments to reduce the autofluorescence in the B.cockerelli alimentary canal tissues.Furthermore,we assessed the compatibility of the above‐mentioned treatments with Lso immunolocalization and actin staining using phalloidin.Our results showed that the autofluorescence in the alimentary canal was reduced by irradiation,H2O2,or SBB treatments.The compatibility assays indicated that irradiation and H2O2 treatment both greatly reduced the fluorescent signal associated with Lso and actin.However,the SBB incubation preserved those target signals,while efficiently eliminating autofluorescence in the psyllid alimentary canal.Therefore,herein we propose a robust method for reducing the autofluorescence in the B.cockerelli alimentary canal with SBB treatment,which may improve the use of immunofluorescence labeling in this organism.This method may also have a wide range of uses by reducing the autofluorescence in other arthropod species.
文摘Xyloglucans in the non-lignified primary cell walls of different species of monocotyledons have diverse struc- tures, with widely varying proportions of oligosaccharide units that contain fucosylated side chains （F side chains）. To determine whether fucosylated xyloglucans occur in all non-lignified walls in a range of monocotyledon species, we used immunofluorescence microscopy with the monoclonal antibody CCRC-M1. The epitope of this antibody, α-L-FUCp-（1 →2）- β-D-Galp, occurs in F side chains. In most non-commelinid monocotyledons, the epitope was found in all non-lignified walls. A similar distribution was found in the palm Phoenix canariensis, which is a member of the basal commelinid order Arecales. However, in the other commelinid orders Zingiberales, Commelinales, and Poales, the occurrence of the epitope was restricted, sometimes occurring in only the phloem walls, but often also in walls of other cell types including stomatal guard and subsidiary cells and raphide idioblasts. No epitope was found in the walls of the commelinids Tradescantia virginiana （Commelinaceae, Commelinales） and Zea mays （Poaceae, Poales）, but it occurred in the phloem walls of two other Poaceae species, Lolium multiflorum and L. perenne. The distribution of the epitope is discussed in relation to xyloglucan structures in the different taxa. However, the functional significance of the restricted distributions is unknown.
文摘Erratum to:J Huazhong Univ Sci Technol[Med Sci]36(4):548–553,2016 https://doi.org/10.1007/s11596-016-1623-6 In the originally published article(https://doi.org/10.1007/s11596-016-1623-6),the immunofluorescence images in shRNA group in Fig.3 were accidentally used rather than the final,formal experiments.To retain consistency,the entire Fig.3 is replaced here with original images of the experiments.The authors declare that this correction will not affect the conclusion of the study.
基金supported by the National Natural Science Foundation of China(U22A20374,52373273)National High Level Hospital Clinical Research Funding of China-Japan Friendship Hospital(Grant number:2024-NHLHCRF-JBGSWZ-02).
文摘With the deepening of epigenetic research,studies have shown that N6-methyladenosine(m^(6)A)is closely related to the development of rheumatoid arthritis(RA),but the mechanism is still unclear.In the study,we collected synovial tissues from normal controls and patients with osteoarthritis(OA)or RA.The levels of m^(6)A and inflammation were analyzed by immunofluorescence staining and western blotting.The roles of IGF2BP3 in cell proliferation and inflammatory activation were explored using transfection and RNA immunoprecipitation assays.IGF2BP3^(−/−)mice were generated and used to establish an arthritis mouse model by transferring serum from adult arthritis K/BxN mice.We found m^(6)A levels were markedly increased in RA patients and mouse models,and the expression of IGF2BP3 was upregulated in individuals with RA and related to the levels of inflammatory markers.IGF2BP3 played an important part in RA-fibroblast-like synoviocytes(FLS)by promoting cell proliferation,migration,invasion,inflammatory cytokine release and inhibiting autophagy.In addition,IGF2BP3 inhibited autophagy to reduce ROS production,thereby decreasing the inflammatory activation of macrophages.More importantly,RASGRF1-mediated mTORC1 activation played a crucial role in the ability of IGF2BP3 to promote cell proliferation and inflammatory activation.In an arthritis model of IGF2BP3^(−/−)mice,IGF2BP3 knockout inhibited RA-FLS proliferation and inflammatory infiltration,and further ameliorated RA joint injury.Our study revealed an important role for IGF2BP3 in RA progression.The targeted inhibition of IGF2BP3 reduced cell proliferation and inflammatory activation and limited RA development,providing a potential strategy for RA therapy.
基金Supported by Panyu District Science and Technology Plan Project,No.2024-Z04-012.
文摘BACKGROUND Patients with tumors often develop multiple autoantibodies against tumorassociated antigens.Among these,antinuclear antibodies(ANAs)constitute a clinically important group distributed across the nucleus,cytoplasm,and cytoskeleton.Emerging evidence suggests that ANAs are closely associated with the development and progression of various malignancies,including colorectal cancer(CRC).AIM To detect ANA fluorescence patterns in CRC using indirect immunofluorescence(IIF)and investigate their correlation with the disease.METHODS We collected serum samples from patients and healthy controls visiting The Affiliated Panyu Central Hospital of Guangzhou Medical University between May 2023 and March 2024 for analysis.The study included 38 patients with newly diagnosed CRC,43 patients with colorectal polyps(CRP),and 29 healthy controls.Serum ANA expression was assessed by IIF,and fluorescence patterns were recorded for each group.Differences in ANA titers were compared among each group to analyze the differences in serum ANA-positive expression,which were further analyzed to explore the correlation between ANA expression and CRC screening.RESULTS ANA positivity rates were 50.00%in the CRC group,46.51%in the colorectal polyp group,and 6.90%in the healthy control group,with significantly higher rates in the two patient groups compared to the control group(P<0.05).In the CRC group,the most common fluorescence patterns were nuclear speckled(15.79%)and cytoplasmic speckled(15.79%),with titers predominantly low(1:100,28.95%).In the colorectal polyp group,nuclear speckled(18.60%)and nuclear homogeneous(11.63%)were the most frequent,with titers also predominantly low(1:100,37.21%).The distribution of intermediate titers differed significantly among groups(P<0.05).CONCLUSION ANAs are associated with both CRP and CRC and may be useful in early CRC screening.Medium-to-high ANA titers,in particular,should prompt further evaluation for possible CRC correlation.Multiple ANA fluorescence patterns can be detected across all groups,with patients with CRP and CRC showing greater pattern diversity than healthy controls.
基金Supported by Science and Technology Foundation of PLA General Lo-gistics Department(06G138)~~
文摘[Objective] The aim of the present study was to develop a direct immunofluorescence method for the diagnosis of canine distemper （CD） with FITC-conjugated monoclonal antibodies （FITC-McAb）.[ Metbod] The McAb against CDV, designated as CE3, was purified with protein G and labeled with FITC through agitation method. After purification and identification, the optimal working concentration of FITC-labeled CE3 was determined. Then 61 clinical samples of suspected canine distemper were detected by direct immunofluorescence assay. [ Result] The absorption test, blocking test and specificity test showed that the labeled antibody had high specificity and sensitivity, but didn＇t have cross reaction with canine parvovirus （CPV）, canine parainfluenza virus （CPIV）, canine adenovirus （CAV） and rabies virus （RV）. The optimal working concentration was 1：80. The positive rate of clinical suspected samples was 48%. [ Conclusion] The direct immunofluorescence assay developed in this study was rapid, specific and convenient, and had great significance for the early diagnosis of canine distemper.
文摘Background: Diarrheal diseases have globally decreased over the past few decades, yet they remain one of the top three causes of mortality in children under five years, especially in sub-Saharan Africa and Nigeria. Seasonal peaks of diarrheal episodes continue to contribute significantly to childhood mortality in these regions. One of the notable causes of diarrhea in children is parasitic infections, particularly Cryptosporidium, which poses a serious health risk. In Nigeria, the burden of Cryptosporidium diarrhea is under-researched, making it imperative to investigate its prevalence and associated risk factors. Study Objectives: The study aims to determine the prevalence and risk factors associated with Cryptosporidium diarrhea among children aged five years and below in selected health institutions in Abakaliki, South-East Nigeria. Methodology: This cross-sectional study was conducted from January to May 2017, recruiting 200 children under five years with diarrhea from health institutions in Abakaliki. Fecal specimens were analyzed for Cryptosporidium oocysts using light microscopy with modified Ziehl-Neelsen staining and immunofluorescent antibody test (IFAT). Deoxyribonucleic acid (DNA) was extracted from positive samples using QIAmp® DNA stool kit, followed by Polymerase Chain Reaction (PCR) and molecular genotyping. Results: Cryptosporidium was detected in 0.5% (1/200) of children via light microscopy and 6.5% (13/200) via IFAT. All positive samples were confirmed as Cryptosporidium hominis by PCR. The prevalence of infection was significantly higher in children from institutionalized homes (50.0%) compared to monogamous homes (6.2%) (p Conclusion: Cryptosporidium hominis is a notable cause of diarrhea among children in Abakaliki, primarily transmitted through human-to-human contact. The study underscores the need for targeted interventions in childcare institutions to prevent outbreaks. Health authorities should promote breastfeeding and enhance education on hygiene practices in vulnerable populations.
文摘Objective: To study the relationship between Chlamydia pneumoniae (C. pneumoniae) infection and asthma exacerbation. Methods: A prospective study of C. pneumoniae infection was conducted in 75 patients with asthma and 63 patients with respiratory tract infection, and 100 blood donors served as controls. The presence of infection was convinced by the polymerase chain reaction and direct immunofluorescence assay for C. pneumoniae DNA from throat swab specimens and micro-immunofluorescence testing for C. pneu-moniae-specific IgG, IgM and IgA antibodies. Results: Prevalence of specific IgG in asthma patients (81. 3%) was higher than that of the blood donors (68. 0%, P<0. 05) and was not significantly different from respiratory tract infection patients (68. 0%, P>0. 05). The acute C. pneumoniae infection rate of symptomatic asthma patients (59. 4%) was markedly higher than that of respiratory tract infection patients (34. 9% , P<0. 05). The average titer of C. pneumoniae IgG instead of IgA in asthma patients (48. 38±6. 94) was significantly higher than respiratory tract infection patients (24. 70±8. 77, P<0. 05). Other pathogens were identified in 12 of 21 (57. 1%) asthma patients with C. pneumoniae. The symptoms of 7 asthma patients with C. pneumoniae infection were improved through antibiotic treatment. Conclusion: The findings suggest a possible role of C. pneumoniae infection in asthma.
文摘One of the prominent cell cycle related modifications of histone proteins, whose function is correlated with chromosome condensation, is the phosphorylation of histone H3. Wheat (Triticum aestivum L.) mitotic and meiotic cells were analyzed with indirect immunoflurorescence labeling with an antibody recognizing histone H3 phosphorylated at Serine 10 to study the localization of phosphorylated histone H3 at mitosis and meiosis. Our results showed that, during mitotic division, the phosphoryiation of H3 started from early prophase and vanished at telophase, remaining mainly in the pericentromeric regions at metaphase and anaphase. During meiotic division, phosphorylation of H3 initiated at the transition from leptotene to zygotene and remained uniform, along the chromosomes from prophase I until telophase whereas it showed slightly stronger in the pericentromeric regions than along the chromosome arms from metaphase II until Lelophase II The different patterns of H3 phophorylation at mitosis and meiosis in wheat suggested that this evolutionarily conserved post-translational chromatin modification might be involved in more roles besides chromosome condensation.
基金Supported by the National Key Technologies Research and Develop-ment Program of China during the 10th Five-Year Plan Period(2004BA519A05)Technologies Research and Development Program of China during the 10th Five-Year Plan Period in Jiangsu Province(BE2002346).~~
文摘Five monoclonal antibodies（Mabs） to nuclear protein of avain influenza virus（AIV） were developed by syncretizing SP 2/0 and the spleen cells from BALB of mice immuized with H9 subtype AIV. Specificity of these Mabs were identified by immunofluorescent assay（IFA） and enzyme linked immunosorbent assay （ELISA）. These five Mabs which were named as AIV-NP-2C3, AIV-NP-6A5, AIV-NP-3 H9, AIV-NP-7B4, AIV-NP-2H4 could react with all viruses of AIV-H9 strains in tests. The result of Western blotting showed that only the 60 ku protein antigen of AIV-H9 could be recognized by the Mabs but never recognized by New castle disease virus, REV and infectious bursa disease virus. The result of preliminary application showed that avian influenza viruses could be deetected bv Mabs in IFA and ELISA. All these Mabs will probably play important roles in preventing and monitoring avian influenza viruses.
基金Grant-in-aid No. 39970344 and No. 30470798the National Nature Science Foundation, China in 1999 and 2004
文摘AIM: To investigate whether KAI1, as a metastasis suppressor gene, is associated with invasive and metastatic ability of pancreatic cancer cells.METHODS: KAI1 gene was transfected into pancreatic cancer cell line MiaPaCa Ⅱ by liposomes selected with G418. Expression of transfected cells was measured by Western blotting, immunofluorescence and immunocytochemistry. Tumor cell invasion and metastatic ability were detected through gelatinase activity and reconstituted basement membrane (Matrigel) assay. pCMV-KAI1 was directly injected into the heterotopic human pancreatic adenocarcinoma successfully established in the groin of BALB/C nude mice, by subcutaneous injection of MiaPaCa Ⅱ pancreatic cancer cells. The statistical analysis between groups was determined by Student's two tailed t test.RESULTS: By Western blotting, MiaPaCa Ⅱ cells transfected by KAI1 gene indicated KAI1 expression at approximately 29.1 kDa. Cytoplasm staining was positive and uniformly spread in transfected cancer cells, using immunohistochemistry and immunofluorescence. The most obvious difference was present after 30 h (MiaPaca Ⅱ 43.6 ± 9.42, pCMV-MiaPaca Ⅱ 44.8 ± 8.56, pCMV-KAI1-MiaPaca Ⅱ 22.0 ± 4.69, P < 0.05). Gelatinolysis revealed a wider and clearer band of gelatinolytic activity in non-transfected than in transfected cells (MiaPaCa Ⅱ cells 30.8 ± 0.57, transfected cells 28.1 ± 0.65, P < 0.05). In vivo tumor growth rates of KAI1 transfectants with KAI1-Lipofectamine 1.22 ± 0.31 in A group were lower than control 4.61 ± 1.98 and pCMV-KAI 11.67 ± 0.81. Analyses of metastases with and without KAI1 transfection in mice were different in liver and lung between controls 1.62 ± 0.39, 0.45 ± 0.09, pCMV-KAI 1.01 ± 0.27, 0.33 ± 0.09 and KAI1-Lipofectamine 0.99 ± 0.21, 0.30 ± 0.09 respectively (P < 0.05).CONCLUSION: High expression of KAI1 gene was found in transfected MiaPaCa Ⅱ human pancreatic cancer cells with lower metastatic ability. KAI1 gene plays an important role in inhibiting metastasis of pancreatic cancer after direct injection into pancreatic adenocarcinoma. These results show that the suppressed invasion and motor function of pancreatic cancer cells may be a key reason why the KAI1 gene controls pancreatic cancer cell metastasis.
