Journal articles: 'Plaque reduction assay'

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Li, Xinhui, and Haiqiang Chen. "Evaluation of the Porcine Gastric Mucin Binding Assay for High-Pressure-Inactivation Studies Using Murine Norovirus and Tulane Virus." Applied and Environmental Microbiology 81, no. 2 (October 31, 2014): 515–21. http://dx.doi.org/10.1128/aem.02971-14.

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ABSTRACTWe compared the results of high-hydrostatic-pressure (HHP) inactivation of murine norovirus type 1 (MNV-1) and Tulane virus (TV) obtained by a porcine gastric mucin binding assay followed by quantitative reverse transcription-PCR (referred to here as the PGM-MB/PCR assay) and a plaque assay and evaluated HHP inactivation of a human norovirus (HuNoV) genogroup I genotype 1 (GI.1) strain and a HuNoV GII.4 strain by using the PGM-MB/PCR assay. Viruses were treated at different pressure levels for 2 min at 4 or 21°C in culture medium of neutral pH and in culture medium of pH 4 at 21°C. The log reductions of infectious MNV-1 and TV particles caused by HHP were assessed using the PGM-MB/PCR and plaque assays, while the log reductions of HuNoVs were assessed by the PGM-MB/PCR assay only. For TV and MNV-1, the two pressure inactivation curves obtained using the plaque and PGM-MB/PCR assays were almost identical at ≤2-log-reduction levels regardless of the treatment temperature and pH. Further increasing the pressure over the 2-log-reduction level resulted in higher log reductions of TV and MNV-1, as assessed by the plaque assay, but did not increase the log reductions, as assessed by the PGM-MB/PCR assay. HHP treatments could achieve maximum reductions of ∼3 and 3.5 log units for GI.1 and GII.4, respectively, as assessed by the PGM-MB/PCR assay. On the basis of these results, it can reasonably be concluded that the PGM-MB/PCR assay would very likely be able to estimate HHP inactivation of HuNoV at ≤2-log-reduction levels. It would also likely conservatively quantify HHP inactivation of the GI.1 strain at 2- to 3-log-reduction levels and the GII.4 strain at 2- to 3.5-log-reduction levels.

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Arias-Arias, Jorge L., Eugenia Corrales-Aguilar, and Rodrigo A. Mora-Rodríguez. "A Fluorescent Real-Time Plaque Assay Enables Single-Cell Analysis of Virus-Induced Cytopathic Effect by Live-Cell Imaging." Viruses 13, no. 7 (June 22, 2021): 1193. http://dx.doi.org/10.3390/v13071193.

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Conventional plaque assays rely on the use of overlays to restrict viral infection allowing the formation of distinct foci that grow in time as the replication cycle continues leading to countable plaques that are visualized with standard techniques such as crystal violet, neutral red, or immunolabeling. This classical approach takes several days until large enough plaques can be visualized and counted with some variation due to subjectivity in plaque recognition. Since plaques are clonal lesions produced by virus-induced cytopathic effect, we applied DNA fluorescent dyes with differential cell permeability to visualize them by live-cell imaging. We could observe different stages of that cytopathic effect corresponding to an early wave of cells with chromatin-condensation followed by a wave of dead cells with membrane permeabilization within plaques generated by different animal viruses. This approach enables an automated plaque identification using image analysis to increase single plaque resolution compared to crystal violet counterstaining and allows its application to plaque tracking and plaque reduction assays to test compounds for both antiviral and cytotoxic activities. This fluorescent real-time plaque assay sums to those next-generation technologies by combining this robust classical method with modern fluorescence microscopy and image analysis approaches for future applications in virology.

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Naveed, Khalid, Aqeel Javeed, Muhammad Ashraf, Amjad Riaz, Aamir Ghafoor, and Adeel Sattar. "Effect of nabumetone on humoral immune responses in mice." Arquivo Brasileiro de Medicina Veterinária e Zootecnia 72, no. 3 (May 2020): 915–20. http://dx.doi.org/10.1590/1678-4162-11460.

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ABSTRACT Nabumetone is used to reduce the pain and inflammation in rheumatoid arthritis. In the current study, immunomodulatory effect of Nabumetone is investigated in mice. The control group was administered normal saline orally as placebo. Nabumetone was administered orally via gavage in two treatment groups at 14mg/kg.b.w. doses and 28mg/kgb.w., respectively. Haemagglutination (HA) assay, Jerne hemolytic plaque and mice lethality assays were applied. In HA assay, the titer was significantly decreased in Nabumetone treatment groups (P< 0.001). In Jerne hemolytic plaque formation assay, there was a significant reduction (P< 0.001) in number of plaques in Nabumetone treated groups when compared with control. In mice lethality assay, there was a significant difference in mortality ratio of mice in control and Nabumetone treated groups (P< 0.001). Therefore, it is concluded that Nabumetone suppresses the humoral immune response in mice.

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Leary, Jeffry J., Robert Wittrock, Robert T. Sarisky, Adriana Weinberg, and Myron J. Levin. "Susceptibilities of Herpes Simplex Viruses to Penciclovir and Acyclovir in Eight Cell Lines." Antimicrobial Agents and Chemotherapy 46, no. 3 (March 2002): 762–68. http://dx.doi.org/10.1128/aac.46.3.762-768.2002.

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ABSTRACT The commonly used antiviral drugs acyclovir (ACV) and penciclovir (PCV) possess similarly potent antiviral activities in vivo against herpes simplex virus (HSV). Assay methods for sensitivity to ACV are not necessarily transferable to PCV, even though the two drugs have similar in vivo potencies and mechanisms of action. We determined by plaque reduction assay the relative activities of ACV and PCV against five laboratory-adapted strains of HSV types 1 and 2 (including sensitive and resistant strains) in seven human cell lines and one nonhuman primate cell line. Seven characteristics were used to evaluate the cell lines. All cell lines were similar in their plating efficiencies and abilities to discriminate between sensitive and resistant HSV isolates. Vero and MRC-5 cells yielded the most discordant 50% inhibitory concentrations (IC50s) for the two HSV types, while Vero and WI-38 VA-13 cells yielded large differences in the IC50s of ACV and PCV. The limited life spans and poor plaque morphologies of the fibroblast lines were undesirable characteristics. Among the transformed cell lines producing well-defined plaques, A549 cells provided the best concordance between IC50s for the two HSV types and two antiherpes drugs. Comparison experiments with a yield reduction format indicated that the use of assays of this type might allow some of the cell-specific properties observed in plaque reduction assays to be avoided.

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McSharry, James M., Nell S. Lurain, George L. Drusano, Alan Landay, Jody Manischewitz, Mostafa Nokta, Maurice O’Gorman, et al. "Flow Cytometric Determination of Ganciclovir Susceptibilities of Human Cytomegalovirus Clinical Isolates." Journal of Clinical Microbiology 36, no. 4 (1998): 958–64. http://dx.doi.org/10.1128/jcm.36.4.958-964.1998.

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A flow cytometric assay has been developed for the measurement of susceptibilities to ganciclovir of laboratory strains and clinical isolates of human cytomegalovirus (HCMV). The assay uses fluorochrome-labeled monoclonal antibodies to HCMV immediate-early and late antigens to identify HCMV-infected cells and flow cytometry to detect and quantitate the number of antigen-positive cells. By this assay, the 50 and 90% inhibitory concentrations (IC50 and IC90, respectively) of ganciclovir for the AD169 strain of HCMV were 1.7 and 9.2 μM, respectively, and the IC50 for the ganciclovir-resistant D6/3/1 derivative of the AD169 strain was greater than 12 μM. The ganciclovir susceptibilities of 17 HCMV clinical isolates were also determined by flow cytometric analysis of the effect of ganciclovir on late-antigen synthesis in HCMV-infected cells. The average IC50 of ganciclovir for drug-sensitive HCMV clinical isolates was 3.79 μM (±2.60). The plaque-reduction assay for these clinical isolates yielded an average IC50of 2.80 μM (±1.46). Comparison of the results of the flow cytometry assays with those obtained from the plaque-reduction assays demonstrated acceptable bias and precision. Flow cytometric and plaque-reduction analysis of cells infected with ganciclovir-resistant clinical isolates failed to show a reduction in the percentage of late-antigen-positive cells or PFU, even at 96 μM ganciclovir. The flow cytometric assay for determining ganciclovir susceptibility of HCMV is quantitative, and objective, and potentially automatable, and its results are reproducible among laboratories.

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de Graaf, Miranda, Sander Herfst, Eefje J. A. Schrauwen, Bernadette G. van den Hoogen, Albert D. M. E. Osterhaus, and Ron A. M. Fouchier. "An improved plaque reduction virus neutralization assay for human metapneumovirus." Journal of Virological Methods 143, no. 2 (August 2007): 169–74. http://dx.doi.org/10.1016/j.jviromet.2007.03.005.

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Stránská, Růžena, Rob Schuurman, David R. Scholl, Joseph A. Jollick, Carl J. Shaw, Caroline Loef, Merjo Polman, and Anton M. van Loon. "ELVIRA HSV, a Yield Reduction Assay for Rapid Herpes Simplex Virus Susceptibility Testing." Antimicrobial Agents and Chemotherapy 48, no. 6 (June 2004): 2331–33. http://dx.doi.org/10.1128/aac.48.6.2331-2333.2004.

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ABSTRACT A colorimetric yield reduction assay, ELVIRA (enzyme-linked virus inhibitor reporter assay) HSV, was developed to determine the antiviral drug susceptibilities of herpes simplex virus (HSV). It uses an HSV-inducible reporter cell line. This simple and rapid assay has an objective readout, low inoculum size, and good reproducibility. The results correlate well with those of the plaque reduction assay.

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Haralambieva, Iana H., Inna G. Ovsyannikova, Robert A. Vierkant, and Gregory A. Poland. "Development of a Novel Efficient Fluorescence-Based Plaque Reduction Microneutralization Assay for Measles Virus Immunity." Clinical and Vaccine Immunology 15, no. 7 (May 7, 2008): 1054–59. http://dx.doi.org/10.1128/cvi.00008-08.

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ABSTRACT The measurement of functional measles virus-specific neutralizing antibodies is of considerable interest for vaccine-related research. In this study, we developed and standardized a simple, rapid, highly sensitive, and reproducible fluorescence-based plaque reduction microneutralization (PRMN) assay with visual and automated readout, using a recombinant measles virus engineered to express enhanced green fluorescent protein. The assay is performed in micro format, requires less time to complete (2 versus 4 to 7 days), and is less labor-intensive and less costly than the classical plaque reduction neutralization (PRN) test, widely accepted as the “gold standard” in measles serology. Two available WHO international anti-measles virus standards and one in-house reference serum were used to develop and standardize the new assay. The mean PRMN values from repeated assays were found to be similar to those reported in the literature or assigned to the WHO standards by the classical PRN assay. For validation, we used three groups of low, moderate, and high measles virus vaccine responders’ sera with moderate values of correlation in antibody levels (mIU/ml) between PRMN and the Dade Behring immunoglobulin G enzyme immunoassay (EIA). The PRMN assay was more sensitive at low antibody levels and more informative in terms of protection than this commercial EIA. In conclusion, we have developed and validated a sensitive and high-throughput measles virus-specific PRMN that can be readily used in large population-based measles studies.

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Landry, Marie L., Sylvia Stanat, Karen Biron, Donald Brambilla, William Britt, Janet Jokela, Sunwen Chou, et al. "A Standardized Plaque Reduction Assay for Determination of Drug Susceptibilities of Cytomegalovirus Clinical Isolates." Antimicrobial Agents and Chemotherapy 44, no. 3 (March 1, 2000): 688–92. http://dx.doi.org/10.1128/aac.44.3.688-692.2000.

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ABSTRACT Twelve laboratories collaborated in formulating and testing a standardized plaque reduction assay for cytomegalovirus (CMV) cell-associated clinical isolates. Four characterized and plaque-purified CMV strains, as well as six coded clinical isolates obtained after antiviral therapy, were distributed and tested. Good agreement was obtained for four of the clinical isolates, but a broad distribution of results was obtained for two isolates. Analysis of these results indicates the problems associated with clinical isolates, including the large genetic variability and the highly cell-associated phenotype. This collaborative effort, by addressing these problems, represents a significant step toward the development of a standardized assay.

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Schaack, Jerome, William Y. Ho, Shawna Tolman, Elizabeth Ullyat, Xiaoling Guo, Nina Frank, Paul I. Freimuth, Dick J. Roovers, and John S. Sussenbach. "Construction and Preliminary Characterization of a Library of “Lethal” Preterminal Protein Mutant Adenoviruses." Journal of Virology 73, no. 11 (November 1, 1999): 9599–603. http://dx.doi.org/10.1128/jvi.73.11.9599-9603.1999.

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ABSTRACT Adenoviruses containing lethal in-frame insertion mutant alleles of the preterminal protein (pTP) gene were constructed with cell lines that express pTP. Thirty in-frame insertion mutant alleles, including 26 alleles previously characterized as lethal and 4 newly constructed mutant alleles, were introduced into the viral chromosome in place of the wild-type pTP gene. The viruses were tested for ability to form plaques at 37°C in HeLa-pTP cells and at 32°C and 39.5°C in HeLa cells. Two of the newly constructed viruses exhibited temperature sensitivity for plaque formation, one virus did not form plaques in the absence of complementation, seven additional mutants exhibited a greater than 10-fold reduction in plaque formation in the absence of complementation, and another eight mutants exhibited stronger phenotypes than did previously characterized in-frame insertion mutants in the plaque assay. These mutant viruses offer promise for analysis of pTP functions.

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McSharry, James J., Nell S. Lurain, George L. Drusano, Alan L. Landay, Mostafa Notka, Maurice R. G. O’Gorman, Adriana Weinberg, Howard M. Shapiro, Patricia S. Reichelderfer, and Clyde S. Crumpacker. "Rapid Ganciclovir Susceptibility Assay Using Flow Cytometry for Human Cytomegalovirus Clinical Isolates." Antimicrobial Agents and Chemotherapy 42, no. 9 (September 1, 1998): 2326–31. http://dx.doi.org/10.1128/aac.42.9.2326.

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ABSTRACT Rapid, quantitative, and objective determination of the susceptibilities of human cytomegalovirus (HCMV) clinical isolates to ganciclovir has been assessed by an assay that uses a fluorochrome-labeled monoclonal antibody to an HCMV immediate-early antigen and flow cytometry. Analysis of the ganciclovir susceptibilities of 25 phenotypically characterized clinical isolates by flow cytometry demonstrated that the 50% inhibitory concentrations (IC50s) of ganciclovir for 19 of the isolates were between 1.14 and 6.66 μM, with a mean of 4.32 μM (±1.93) (sensitive; IC50 less than 7 μM), the IC50s for 2 isolates were 8.48 and 9.79 μM (partially resistant), and the IC50s for 4 isolates were greater than 96 μM (resistant). Comparative analysis of the drug susceptibilities of these clinical isolates by the plaque reduction assay gave IC50s of less than 6 μM, with a mean of 2.88 μM (±1.40) for the 19 drug-sensitive isolates, IC50s of 6 to 8 μM for the partially resistant isolates, and IC50s of greater than 12 μM for the four resistant clinical isolates. Comparison of the IC50s for the drug-susceptible and partially resistant clinical isolates obtained by the flow cytometry assay with the IC50s obtained by the plaque reduction assay showed an acceptable correlation (r 2 = 0.473; P = 0.001), suggesting that the flow cytometry assay could substitute for the more labor-intensive, subjective, and time-consuming plaque reduction assay.

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RECKER, JORDAN D., and XINHUI LI. "Evaluation of Copper Alloy Surfaces for Inactivation of Tulane Virus and Human Noroviruses." Journal of Food Protection 83, no. 10 (September 25, 2020): 1782–88. http://dx.doi.org/10.4315/0362-028x.jfp-19-410.

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ABSTRACT This study evaluated the efficacy of copper alloy surfaces for inactivation of Tulane virus (TV), assessed by plaque assay and porcine gastric mucin–conjugated magnetic bead (PGM-MB) binding assay, followed by quantitative reverse transcription PCR (PGM-MB–RT-qPCR assay). In addition, the efficacy of a copper surface for inactivation of human norovirus (HuNoV) GII.4 Sydney and GI.3B Potsdam strains was evaluated by PGM-MB–RT-qPCR assay. Results of time-dependent inactivation of viruses on copper, bronze, and brass coupons revealed that 15 min of surface treatments of each of the copper and copper alloys achieved >4-log reduction of purified TV, as assessed by plaque assay, while up to 20 min of copper alloy surface treatments only achieved ∼2-log reduction, as assessed by PGM-MB–RT-qPCR assay. As assessed by PGM-MB–RT-qPCR assay, 10 min of copper surface treatments achieved reductions of 3 and 4 log units for HuNoVs GII.4 Sydney and GI.3B Potsdam, respectively. Results from this study suggest that even though PGM-MB–RT-qPCR assay underestimated the efficacy of copper alloy surface inactivation of TV, copper alloy surfaces were able to effectively inactivate TV and HuNoVs. Therefore, copper alloys can be used as a preventive measure to prevent HuNoV infection and are an effective surface treatment for HuNoVs. HIGHLIGHTS

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Yepes-Perez, Andres F., Oscar Herrera-Calderón, Cristian A. Oliveros, Lizdany Flórez-Álvarez, María I. Zapata-Cardona, Lina Yepes, Wbeimar Aguilar-Jimenez, María T. Rugeles, and Wildeman Zapata. "The Hydroalcoholic Extract of Uncaria tomentosa (Cat’s Claw) Inhibits the Infection of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) In Vitro." Evidence-Based Complementary and Alternative Medicine 2021 (February 24, 2021): 1–11. http://dx.doi.org/10.1155/2021/6679761.

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The coronavirus disease 2019 (COVID-19) has become a serious problem for public health since it was identified in the province of Wuhan (China) and spread around the world producing high mortality rates and economic losses. Nowadays, the WHO recognizes traditional, complementary, and alternative medicine for treating COVID-19 symptoms. Therefore, we investigated the antiviral potential of the hydroalcoholic extract of Uncaria tomentosa stem bark from Peru against SARS-CoV-2 in vitro. The antiviral activity of U. tomentosa against SARS-CoV-2 in vitro was assessed in Vero E6 cells using cytopathic effect (CPE) and plaque reduction assay. After 48 h of treatment, U. tomentosa showed an inhibition of 92.7% of SARS-CoV-2 at 25.0 μg/mL ( p < 0.0001 ) by plaque reduction assay on Vero E6 cells. In addition, U. tomentosa induced a reduction of 98.6% ( p = 0.02 ) and 92.7% ( p = 0.03 ) in the CPE caused by SARS-CoV-2 on Vero E6 cells at 25 μg/mL and 12.5 μg/mL, respectively. The EC50 calculated for the U. tomentosa extract by plaque reduction assay was 6.6 μg/mL (4.89–8.85 μg/mL) for a selectivity index of 4.1. The EC50 calculated for the U. tomentosa extract by TCID50 assay was 2.57 μg/mL (1.05–3.75 μg/mL) for a selectivity index of 10.54. These results showed that U. tomentosa, known as cat's claw, has an antiviral effect against SARS-CoV-2, which was observed as a reduction in the viral titer and CPE after 48 h of treatment on Vero E6 cells. Therefore, we hypothesized that U. tomentosa stem bark could be promising in the development of new therapeutic strategies against SARS-CoV-2.

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Lewis, Robert B., Debbie S. Matzke, Thomas B. Albrecht, and Richard B. Pollard. "Assessment of the Presence of Cytomegalovirus-Neutralizing Antibody by a Plaque-Reduction Assay." Clinical Infectious Diseases 8, Supplement_4 (July 1, 1986): S434—S438. http://dx.doi.org/10.1093/clinids/8.supplement_4.s434.

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Ward, R. L., A. Z. Kapikian, K. M. Goldberg, D. R. Knowlton, M. W. Watson, and R. Rappaport. "Serum rotavirus neutralizing-antibody titers compared by plaque reduction and enzyme-linked immunosorbent assay-based neutralization assays." Journal of clinical microbiology 34, no. 4 (1996): 983–85. http://dx.doi.org/10.1128/jcm.34.4.983-985.1996.

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Hour, Mann-Jen, Su-Hua Huang, Ching-Yao Chang, Yen-Kuan Lin, Ching-Ying Wang, Yuan-Shiun Chang, and Cheng-Wen Lin. "Baicalein, Ethyl Acetate, and Chloroform Extracts ofScutellaria baicalensisInhibit the Neuraminidase Activity of Pandemic 2009 H1N1 and Seasonal Influenza A Viruses." Evidence-Based Complementary and Alternative Medicine 2013 (2013): 1–11. http://dx.doi.org/10.1155/2013/750803.

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This study rated antiviral activity ofScutellaria baicalensisGeorgi (S. baicalensis) extracts against influenza A virus subtypes, for example, pandemic 2009 H1N1, seasonal H1N1 and H3N2. Ethyl acetate (EtOAc) and chloroform extracts inhibitedin vitroneuraminidase (NA) enzymatic activity and viral replication more than methanol (MeOH) extract. EtOAc extract demonstrated NA inhibition IC50values ranging from 73.16 to 487.40 μg/mL and plaque reduction IC50values ranging from 23.7 to 27.4 μg/mL. Chloroform extract showed antiviral activities with plaque reduction IC50values ranging from 14.16 to 41.49 μg/mL Time-of-addition assay indicated that EtOAc and chloroform extracts also significantly inhibited virus yields after infection. HPLC analysis demonstrated that baicalin was dominant in the MeOH extract; baicalein and chrysin were rich in the EtOAc and chloroform extracts. Molecular simulation revealed baicalein hydrogen bonding with Glu277 as well as hydrophobic and Van der Waals interactions with Ile222, Arg224, Ser246, and Tyr347 in NA1 active sites of NA1. Baicalein inhibited in vitro replication of influenza A viruses pandemic 2009 H1N1 (IC50 = 0.018 μM) and seasonal 2007 H1N1 using plaque reduction assays. A combination of low-dose baicalein with other anti-influenza agents could be applicable for development of alternative remedies treating influenza A virus infection.

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Barnett, J. M., A. Cadman, D. Gor, M. Dempsey, M. Walters, A. Candlin, M. Tisdale, et al. "Zanamivir Susceptibility Monitoring and Characterization of Influenza Virus Clinical Isolates Obtained during Phase II Clinical Efficacy Studies." Antimicrobial Agents and Chemotherapy 44, no. 1 (January 1, 2000): 78–87. http://dx.doi.org/10.1128/aac.44.1.78-87.2000.

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ABSTRACT Zanamivir is a highly selective neuraminidase (NA) inhibitor with demonstrated clinical efficacy against influenza A and B virus infections. In phase II clinical efficacy trials (NAIB2005 and NAIB2008), virological substudies showed mean reductions in virus shedding after 24 h of treatment of 1.5 to 2.0 log1050% tissue culture infective doses compared to a placebo, with no reemergence of virus after the completion of therapy. Paired isolates (n = 41) obtained before and during therapy with zanamivir demonstrated no shifts in susceptibility to zanamivir when measured by NA assays, although for a few isolates NA activity was too low to evaluate. In plaque reduction assays in MDCK cells, the susceptibility of isolates to zanamivir was extremely variable even at baseline and did not correlate with the speed of resolution of virus shedding. Isolates with apparent limited susceptibility to zanamivir by plaque reduction proved highly susceptible in vivo in the ferret model. Further sequence analysis of paired isolates revealed no changes in the hemagglutinin and NA genes in the majority of isolates. The few changes observed were all natural variants. No amino acid changes that had previously been identified in vitro as being involved with reduced susceptibility to zanamivir were observed. These studies highlighted problems associated with monitoring susceptibility to NA inhibitors in the clinic, in that no reliable cell-based assay is available. At present the NA assay is the best available predictor of susceptibility to NA inhibitors in vivo, as measured in the validated ferret model of infection.

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Chutkowski, Christine, Betty Olson, Ann McDonough, James Mahoney, and James J. McSharry. "Use of a Single Monoclonal Antibody To Determine the Susceptibilities of Herpes Simplex Virus Type 1 and Type 2 Clinical Isolates to Acyclovir." Clinical and Vaccine Immunology 9, no. 6 (November 2002): 1379–81. http://dx.doi.org/10.1128/cdli.9.6.1379-1381.2002.

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ABSTRACT This report describes a flow cytometry drug susceptibility assay that uses a single fluorochrome-labeled monoclonal antibody to determine the acyclovir susceptibilities of herpes simplex virus (HSV) type 1 or type 2 clinical isolates. This assay yields 50% effective doses (drug concentrations that reduce the number of antigen-positive cells by 50%) for HSV clinical isolates that are equivalent to those obtained with the plaque reduction assay.

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Safrin, S., E. Palacios, and B. J. Leahy. "Comparative evaluation of microplate enzyme-linked immunosorbent assay versus plaque reduction assay for antiviral susceptibility testing of herpes simplex virus isolates." Antimicrobial Agents and Chemotherapy 40, no. 4 (April 1996): 1017–19. http://dx.doi.org/10.1128/aac.40.4.1017.

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We tested the antiviral susceptibilities of 30 clinical isolates of herpes simplex virus using the microplate in situ enzyme-linked immunosorbent assay (MISE) and the plaque reduction assay (PRA). There was concordance for 26 of 30 acyclovir results and all 30 foscarnet results. MISE and PRA results each predicted the response to acyclovir in 12 of 14 instances and the response to foscarnet in 8 instances. MISE is more rapid than PRA, has an objective endpoint, and correlates well with the clinical response to therapy.

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Kim, Ye Won, Hyun Ju You, Soyoung Lee, Bomi Kim, Do Kyung Kim, Joo-Bong Choi, Ji-Ah Kim, et al. "Inactivation of Norovirus by Lemongrass Essential Oil Using a Norovirus Surrogate System." Journal of Food Protection 80, no. 8 (July 12, 2017): 1293–302. http://dx.doi.org/10.4315/0362-028x.jfp-16-162.

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ABSTRACT This study investigated the effect of lemongrass essential oil (LGEO) on the infectivity and viral replication of norovirus. Murine norovirus 1 (MNV-1), a surrogate of human norovirus, was preincubated with LGEO and then used to infect RAW 264.7 cells in a plaque reduction assay. LGEO exhibited a significant reduction in MNV-1 plaque formation in both time- and dose-dependent manners. The quantification of viral genome by quantitative real-time PCR showed similar results in line with those of the plaque reduction assay. It was revealed that citral, a single compound in LGEO, showed dramatic reduction in MNV-1 infectivity (−73.09% when using a treatment of 0.02%, v/v). The inhibitory activity of LGEO on viral replication was further investigated in HG23 cells that harbored a human norovirus replicon. LGEO treatment significantly reduced viral replication in HG23 cells, which suggests that LGEO may have dual inhibitory activities that inactivate viral coat proteins required for viral infection and suppress norovirus genome replication in host cells. In animal experiments, oral administration of murine norovirus preincubated with LGEO significantly suppressed virus infectivity in vivo. Collectively, these results suggest that LGEO, in particular the LGEO component citral, inactivates the norovirus and its subsequent replication in host cells. Thus, LGEO shows promise as a method of inhibiting norovirus within the food industry.

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Cohen, B. J., R. P. Parry, D. Doblas, D. Samuel, L. Warrener, N. Andrews, and D. Brown. "Measles immunity testing: Comparison of two measles IgG ELISAs with plaque reduction neutralisation assay." Journal of Virological Methods 131, no. 2 (February 2006): 209–12. http://dx.doi.org/10.1016/j.jviromet.2005.08.001.

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Sittek, Lisa-Marie, Thomas Michael Schmidts, and Peggy Schlupp. "Ingredients Acting as a Physical Barrier for the Prevention and Treatment of the Rhinovirus Infection." Applied Sciences 10, no. 18 (September 18, 2020): 6511. http://dx.doi.org/10.3390/app10186511.

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Although the common cold, usually caused by human rhinoviruses, is responsible for enormous damage to the economy and health every year, there are hardly any treatment options or prophylaxis for rhinovirus infections. In this work, the potential of a hydrogel complex, based on polymers and an aqueous extract of Icelandic moss in isla® medic lozenges (Engelhard Arzneimittel, Niederdorfelden, Germany), and two other hydrogels (based on solely xanthan gum or sodium hyaluronate) are investigated for the first time in order to prevent and treat rhinovirus infections. By means of rheological investigations, we demonstrate that isla® medic and containing polymers cause artificial saliva to thicken. Additionally, we demonstrate that the thickening results in the formation of a physical diffusion barrier, which leads to a significant reduction in plaques in a preventive plaque assay with rhinovirus 14 (RV-14). Furthermore, it is shown in a curative plaque assay that the hydrogel complex also has a curative effect on rhinovirus infections in that it reduces the spread of the RV-14-infection on H1-HeLa cell monolayers. Overall, polymer-based hydrogels and related products, such as isla® medic, could contribute to the prevention and treatment of rhinovirus infections.

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Christensen, Ekaterina, and Mette Myrmel. "Coagulant residues’ influence on virus enumeration as shown in a study on virus removal using aluminium, zirconium and chitosan." Journal of Water and Health 16, no. 4 (May 17, 2018): 600–613. http://dx.doi.org/10.2166/wh.2018.028.

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Abstract Research on microorganism reduction by physicochemical water treatment is often carried out under the assumption that the microbiological enumeration techniques are not affected by the presence of coagulants. Data presented here indicate that bacteriophage enumeration by plaque assay and RT-qPCR (reverse transcription quantitative polymerase chain reaction) can be affected by these water treatment chemicals. Treatment of water samples with an alkaline protein-rich solution prior to plaque assay and optimization of RNA extraction for RT-qPCR were implemented to minimize the interference. The improved procedures were used in order to investigate reduction of three viral pathogens and the MS2 model virus in the presence of three coagulants. A conventional aluminium coagulant was compared to alternative agents (zirconium and chitosan) in a coagulation-filtration system. The highest virus reduction, i.e., 99.9–99.99%, was provided by chitosan, while aluminium and zirconium reduced virus by 99.9% in colour-rich water and by 90% in water with less colour, implying an effect of coagulant type and raw water quality on virus reduction. Although charge characteristics of viruses were associated with virus reduction, the results reveal that the MS2 phage is a suitable model for aggregation and retention of the selected pathogens.

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Payment, Pierre, and Michel Trudel. "Influence of inoculum size, incubation temperature, and cell culture density on virus detection in environmental samples." Canadian Journal of Microbiology 31, no. 11 (November 1, 1985): 977–80. http://dx.doi.org/10.1139/m85-185.

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The influence of inoculum size and cell culture density on virus titer by cytopathic effect or plaque assay was studied using poliovirus type 1 and BGM (Buffalo green monkey) cells as a model for this evaluation. With a plaque assay system, a linear relationship was observed for an inoculum size of up 1 mL/25 cm2; a marked decrease in the number of plaques was observed when over 1 mL of sample was inoculated on this surface area. Cell culture density also affected virus titer; maximal titers were observed when cells were seeded at 25 000 to 75 000 cells/mL and incubated for 6 days before infection with the virus. Viral density, evaluated as most-probable-number and measured by cytopathic effect under liquid overlay, revealed that the viral titer was similar up to 1 mL inoculum and increased only when over 1 mL was inoculated. Cell density had no significant effect on the viral titer measured by the most-probable-number method and cytopathic effect. Inactivation of inoculum due to an incubation temperature of 37 °C for a short period was shown to be minimal for poliovirus type 1, reovirus type 2, coxsackievirus B-5, and the simian rotavirus SA-11. Longer inactivation time led to a 2 logs reduction of the infectious titer of coxsackievirus B-5 (in 48 h) while the other viruses showed a significant reduction in titer only after 96 h.

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Morens, D. M., S. B. Halstead, P. M. Repik, R. Putvatana, and N. Raybourne. "Simplified plaque reduction neutralization assay for dengue viruses by semimicro methods in BHK-21 cells: comparison of the BHK suspension test with standard plaque reduction neutralization." Journal of Clinical Microbiology 22, no. 2 (1985): 250–54. http://dx.doi.org/10.1128/jcm.22.2.250-254.1985.

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Pujol, CA, JM Estevez, MJ Carlucci, M. Ciancia, AS Cerezo, and EB Damonte. "Novel DL-Galactan Hybrids from the Red Seaweed Gymnogongrus Torulosus are Potent Inhibitors of Herpes Simplex Virus and Dengue Virus." Antiviral Chemistry and Chemotherapy 13, no. 2 (April 2002): 83–89. http://dx.doi.org/10.1177/095632020201300202.

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A novel series of DL-galactan hybrids extracted from the red seaweed Gymnogongrus torulosus, was evaluated for its in vitro antiviral properties against herpes simplex virus type 2 (HSV-2) and dengue virus 2 (DEN-2). These compounds were very active against both viruses with inhibitory concentration 50% (IC50) values in the range 0.6–16 μg/ml for HSV-2 and 0.19–1.7 μg/ml for DEN-2, respectively, as determined in a virus plaque reduction assay in Vero cells. The DL-galactans lacked of cytotoxic effects, on stationary as well as on actively dividing cells, and anticoagulant properties. Some of the compounds showed a variable level of direct inactivating effect on both virions, with virucidal concentration 50% values exceeding the IC50s obtained by plaque reduction assay. Full inhibitory activity was achieved when the galactans were present during virus adsorption period, suggesting that the mode of action of these compounds is an interference in the binding of the surface envelope glycoprotein with the cell receptor.

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Cock, Ian, and FR Kalt. "A modified MS2 bacteriophage plaque reduction assay for the rapid screening of antiviral plant extracts." Pharmacognosy Research 2, no. 4 (2010): 221. http://dx.doi.org/10.4103/0974-8490.69108.

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Yokobata, Kathy E., John M. Jordan, Orvrlle Chapman, and Curt Krerl. "DEVELOPMENT OF A PLAQUE REDUCTION ASSAY AND APPLICATION TO THE STUDY OF PSORALEN-DAMAGED DNA." Photochemistry and Photobiology 43, no. 4 (April 1986): 391–401. http://dx.doi.org/10.1111/j.1751-1097.1986.tb05620.x.

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Lee, Hee-Jung, Kyung-Il Min, Ki Hoon Park, Hyo Jung Choi, Min-Kyoung Kim, Chi-Young Ahn, Young-Jin Hong, and Young Bong Kim. "Comparison of JEV neutralization assay using pseudotyped JEV with the conventional plaque-reduction neutralization test." Journal of Microbiology 52, no. 5 (March 7, 2014): 435–40. http://dx.doi.org/10.1007/s12275-014-3529-y.

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Xie, Xuping, Marisa C. Nielsen, Antonio E. Muruato, Camila R. Fontes-Garfias, and Ping Ren. "Evaluation of a SARS-CoV-2 lateral flow assay using the plaque reduction neutralization test." Diagnostic Microbiology and Infectious Disease 99, no. 2 (February 2021): 115248. http://dx.doi.org/10.1016/j.diagmicrobio.2020.115248.

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Yuan, Qiuju, Jian Yang, Yan-Fang Xian, Rong Liu, Chun W. Chan, Wutian Wu, and Zhi-Xiu Lin. "The Effect of Spinal Cord Injury on Beta-Amyloid Plaque Pathology in TgCRND8 Mouse Model of Alzheimer’s Disease." Current Alzheimer Research 17, no. 6 (October 7, 2020): 576–86. http://dx.doi.org/10.2174/1567205017666200807191447.

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Background: The accumulation and aggregation of Aβ as amyloid plaques, the hallmark pathology of the Alzheimer.s disease, has been found in other neurological disorders, such as traumatic brain injury. The axonal injury may contribute to the formation of Aβ plaques. Studies to date have focused on the brain, with no investigations of spinal cord, although brain and cord share the same cellular components. Objective: We utilized a spinal cord transection model to examine whether spinal cord injury acutely induced the onset or promote the progression of Aβ plaque 3 days after injury in TgCRND8 transgenic model of AD. Methods: Spinal cord transection was performed in TgCRND8 mice and its littermate control wild type mice at the age of 3 and 20 months. Immunohistochemical reactions/ELISA assay were used to determine the extent of axonal damage and occurrence/alteration of Aβ plaques or levels of Aβ at different ages in the spinal cord of TgCRND8 mice. Results: After injury, widespread axonal pathology indicated by intra-axonal co-accumulations of APP and its product, Aβ, was observed in perilesional region of the spinal cord in the TgCRND8 mice at the age of 3 and 20 months, as compared to age-matched non-TgCRND8 mice. However, no Aβ plaques were found in the TgCRND8 mice at the age of 3 months. The 20-month-old TgCRND8 mice with established amyloidosis in spinal cord had a reduction rather than increase in plaque burden at the lesion site compared to the tissue adjacent to the injured area and corresponding area in sham mice following spinal cord transection. The lesion site of spinal cord area was occupied by CD68 positive macrophages/ activated microglia in injured mice compared to sham animals. These results indicate that spinal cord injury does not induce the acute onset and progression of Aβ plaque deposition in the spinal cord of TgCRND8 mice. Conversely, it induces the regression of Aβ plaque deposition in TgCRND8 mice. Conclusion: The findings underscore the dependence of traumatic axonal injury in governing acute Aβ plaque formation and provide evidence that Aβ plaque pathology may not play a role in secondary injury cascades following spinal cord injury.

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Greengard, Olga, Natalia Poltoratskaia, Evgenia Leikina, Joshua Zimmerberg, and Anne Moscona. "The Anti-Influenza Virus Agent 4-GU-DANA (Zanamivir) Inhibits Cell Fusion Mediated by Human Parainfluenza Virus and Influenza Virus HA." Journal of Virology 74, no. 23 (December 1, 2000): 11108–14. http://dx.doi.org/10.1128/jvi.74.23.11108-11114.2000.

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ABSTRACT 4-GU-DANA (zanamivir) (as well as DANA and 4-AM-DANA) was found to inhibit the neuraminidase activity of human parainfluenza virus type 3 (HPF3). The viral neuraminidase activity is attributable to hemagglutinin-neuraminidase (HN), an envelope protein essential for viral attachment and for fusion mediated by the other envelope protein, F. While there is no evidence that HN's neuraminidase activity is essential for receptor binding and syncytium formation, we found that 4-GU-DANA prevented hemadsorption and fusion of persistently infected cells with uninfected cells. In plaque assays, 4-GU-DANA reduced the number (but not the area) of plaques if present only during the adsorption period and reduced plaque area (but not number) if added only after the 90-min adsorption period. 4-GU-DANA also reduced the area of plaques formed by a neuraminidase-deficient variant, confirming that its interference with cell-cell fusion is unrelated to inhibition of neuraminidase activity. The order-of-magnitude lower 50% inhibitory concentrations of 4-GU-DANA (and also DANA and 4-AM-DANA) for plaque area reduction and for inhibition in the fusion assay than for reducing plaque number or blocking hemadsorption indicate the particular efficacy of these sialic acid analogs in interfering with cell-cell fusion. In cell lines expressing influenza virus hemagglutinin (HA) as the only viral protein, we found that 4-GU-DANA had no effect on hemadsorption but did inhibit HA2b-red blood cell fusion, as judged by both lipid mixing and content mixing. Thus, 4-GU-DANA can interfere with both influenza virus- and HPF3-mediated fusion. The results indicate that (i) in HPF3, 4-GU-DANA and its analogs have an affinity not only for the neuraminidase active site of HN but also for sites important for receptor binding and cell fusion and (ii) sialic acid-based inhibitors of influenza virus neuraminidase can also exert a direct, negative effect on the fusogenic function of the other envelope protein, HA.

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Nasser, A. M., Y. Tchorch, and B. Fattal. "Validity of serological methods (ELISA) for detecting infectious viruses in water." Water Science and Technology 31, no. 5-6 (March 1, 1995): 307–10. http://dx.doi.org/10.2166/wst.1995.0632.

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Detection of cultivable enteric viruses in environmental water samples in tissue culture is time consuming and expensive. Moreover, some important enteric viruses grow very slowly (Hepatitis A virus) or do not grow as yet (Norwalk) in tissue culture. Therefore, sensitive serological and molecular methods have been developed to simplify and speed the detection of viruses in environmental samples. This study was conducted to test the reliability of serological methods to monitor the presence of viable viruses in natural waters. The study was performed with poliovirus purified in CsCl gradients and impure virus. Poliovirus 1 either purified or impure was seeded in raw wastewater, groundwater and phosphate buffered saline (PBS) and incubated for 20 days at 4°C, 20°C and 30°C. Virus survival was monitored by a nylon filter A-ELISA and plaque-assay in BGM cells. In all water samples at 4°C, no die-off was observed neither by A-ELISA nor by plaque-assay. In wastewater and groundwater at 20°C and 30°C, greater die-off was observed with A-ELISA than with plaque-assay. Purified poliovirus was undetectable by the A-ELISA after two days of incubation at 20°C in wastewater and groundwater, whereas under the same conditions, only 2log10 reduction were observed in the titer of poliovirus 1. The data of this study have shown that in all cases, a positive test by A-ELISA was also positive by the plaque-assay. Therefore, a positive result of A-ELISA indicates the presence of viable virus in natural waters.

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Jacobson, Jennie G., Thomas E. Renau, M. Reza Nassiri, Dominica G. Sweier, Julie M. Breitenbach, Leroy B. Townsend, and John C. Drach. "Nonnucleoside Pyrrolopyrimidines with a Unique Mechanism of Action against Human Cytomegalovirus." Antimicrobial Agents and Chemotherapy 43, no. 8 (August 1, 1999): 1888–94. http://dx.doi.org/10.1128/aac.43.8.1888.

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ABSTRACT Based upon a prior study which evaluated a series of nonnucleoside pyrrolo[2,3-d]pyrimidines as inhibitors of human cytomegalovirus (HCMV), we have selected three active analogs for detailed study. In an HCMV plaque-reduction assay, compounds 828, 951, and 1028 had 50% inhibitory concentrations (IC50s) of 0.4 to 1.0 μM. Similar results were obtained when 828 and 951 were examined by HCMV enzyme-linked immunosorbent assay (IC50s = 1.9 and 0.4 μM, respectively) and when 828 was tested in a viral DNA-DNA hybridization assay (IC50 = 1.3 μM). In yield-reduction assays with a low multiplicity of infection (MOI), all three compounds caused multiple log10 reductions in virus titer, and the activities of these compounds were comparable to the activity of ganciclovir (GCV; IC90 = 0.2 μM). In contrast to the reduction of viral titers by GCV, the reduction of viral titers by 828, 951, and 1028 decreased with increasing MOI. Cytotoxicity in human foreskin fibroblasts and KB cells ranged from 32 to >100 μM. In addition, 828 (the only compound tested) was less toxic against human bone marrow progenitor cells than GCV. Time-of-addition and time-of-removal studies established that the three pyrrolopyrimidines inhibited HCMV replication before GCV had an effect on viral DNA synthesis but after viral adsorption. Compound 828 was equally effective against GCV-sensitive and GCV-resistant HCMV clinical isolates. Combination studies with 828 and GCV showed that the effects of the two compounds on HCMV were additive but not synergistic. Taken together, the data indicate that these pyrrolopyrimidines target a viral protein that is required in an MOI-dependent manner and that is expressed early in the HCMV replication cycle.

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Eltringham, I. J., S. M. Wilson, and F. A. Drobniewski. "Evaluation of a Bacteriophage-Based Assay (Phage Amplified Biologically Assay) as a Rapid Screen for Resistance to Isoniazid, Ethambutol, Streptomycin, Pyrazinamide, and Ciprofloxacin among Clinical Isolates of Mycobacterium tuberculosis." Journal of Clinical Microbiology 37, no. 11 (1999): 3528–32. http://dx.doi.org/10.1128/jcm.37.11.3528-3532.1999.

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Rapid molecular assays for the detection of mutations associated with rifampin resistance in Mycobacterium tuberculosis are commercially available. However, they are complex and expensive and have predictive values of 90 to 95%. Molecular assays for other drugs are less predictive of resistance. Ideally, assays based on phenotypic markers should be used for susceptibility testing, but these can take weeks to complete. We previously described a rapid phenotypic assay, the phage amplified biologically (PhaB) assay, for the rapid determination of rifampin and isoniazid susceptibility in clinical isolates of M. tuberculosis. In this study, we extended the assay to the study of ethambutol, pyrazinamide, streptomycin, and ciprofloxacin. After the optimization of antibiotic concentrations and incubation conditions, the assay was applied to each drug for a total of 157 isolates. The correlations between the results of the PhaB assay and the resistance ratio method were 94% for isoniazid, 96% for streptomycin, 100% for ciprofloxacin, 88% for ethambutol, and 87% for pyrazinamide. For ciprofloxacin, ethambutol, and pyrazinamide, significantly better correlations were found when a 90% reduction in plaque count was used as the cutoff. Turnaround times for the PhaB assay were 2 to 3 days, compared with 10 days for the resistance ratio method. We believe that this low-cost assay may have widespread applicability for the rapid screening of drug resistance in M. tuberculosis isolates, especially in developing countries.

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Raju, Ramyathilagam, Angeline Divya, Ganesh Rajendran, and James Rufus John. "Analogous assay between green tea mouthwash, listerine mouthwash and chlorhexidine mouthwash in plaque reduction, on orthodontic patients: a randomized cross-over study." International Journal Of Community Medicine And Public Health 4, no. 5 (April 24, 2017): 1429. http://dx.doi.org/10.18203/2394-6040.ijcmph20171751.

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Background: The aim of the study was to compare the efficiency of green tea mouthwash, Listerine mouthwash and Chlorhexidine mouthwash in plaque reduction among orthodontic patients. Methods: The study employed a double blinded, simple randomized, cross over design with a control group consisting of 30 orthodontic patients undergoing fixed appliance therapy. All the subjects were divided into group 1 (Green tea), group 2 (Listerine) and group 3 (Chlorhexidine) as 10 subjects per group. Gingival status was assessed using Sulcus Bleeding Index and plaque accumulation was assessed using Turesky-Gilmore-Glickman modification of Quigley Hein Index. After a relapse period of 15 days, group 1 and 2 were crossed over, however, group 3 remained the same. Indices were again recorded at baseline and 15th day. Results: The mean gingival and plaque score was reduced in all the three groups. However, green tea mouthwash was estimated to have the highest mean difference from 2.17 ± 0.610 at baseline to 1.48 ± 0.474 on the 15th day. Conclusions: Effective use of mouthwashes as supplements for tooth brushing has proved to be beneficial in oral hygiene and maintenance. The findings of this study provide useful insights on the effectiveness of different compositions of mouthwashes.

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Yoakim, C., WW Ogilvie, DR Cameron, C. Chabot, C. Grand-Maître, I. Guse, B. Haché, et al. "Potent β-Lactam Inhibitors of Human Cytomegalovirus Protease." Antiviral Chemistry and Chemotherapy 9, no. 5 (October 1998): 379–87. http://dx.doi.org/10.1177/095632029800900502.

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A series of novel monobactam inhibitors of human cytomegalovirus (HCMV) protease has been described that possess a heterocyclic thiomethyl side chain at C-4. Changes to the heterocycle did not significantly change the inhibitory activity of these compounds in an enzymatic assay, although improvements in solubility and cell culture activity were noted. A number of permutations between C-4 substitutions and N-1 derivatives led to the identification of several β-lactams with antiviral activity in a plaque reduction assay. N-methyl thiotetrazole-containing compounds were found to be the most potent inhibitors in the enzymatic assay.

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Ravault, Stéphanie, Damien Friel, Emmanuel Di Paolo, Adrian Caplanusi, Paul Gillard, Michael Povey, and Stephane Carryn. "Assessment of Mumps Virus-Specific Antibodies: Comparison of Plaque Reduction Neutralization Test and Enzyme-Linked Immunosorbent Assay Estimates." Journal of Infectious Diseases 220, no. 9 (July 11, 2019): 1462–68. http://dx.doi.org/10.1093/infdis/jiz345.

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Abstract Background The plaque reduction neutralization test (PRNT), which measures a subset of immunoglobulin antibodies (functional neutralizing antibodies), and the enzyme-linked immunosorbent assay (ELISA), which measures total immunoglobulin (neutralizing and nonneutralizing antibodies), characterize different aspects of the anti–mumps virus antibody response after vaccination. Methods Data from a recent phase 3 clinical trial (NCT01681992) of 2 measles-mumps-rubella vaccines were used to compare anti-mumps antibody responses measured using an unenhanced PRNT (GSK; seropositivity cutoff and threshold, 2.5 and 4 times the 50% end-point dilution, respectively) with those estimated using an ELISA (thresholds, 5 and 10 ELISA units/mL, respectively). Results Of 3990 initially seronegative samples, 3284 (82.3%) were seropositive after vaccination for anti-mumps antibodies in both assays. The Pearson correlation coefficient for double-positive samples was 0.57, indicative of a moderate correlation. Receiver operating characteristic curve analysis showed that an ELISA threshold of 51.7 ELISA units/mL best corresponded to the PRNT seroresponse threshold. There was no obvious vaccine brand effect on the correlation between assays. Conclusions The moderate correlation between the anti-mumps antibody measurements obtained with PRNT and ELISA reflects different aspects of the serological response. In the absence of a well-defined protective serological threshold, PRNT provides complementary information on the antibody response, whereas ELISA remains a critically useful measurement of vaccine immunogenicity.

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Di Gennaro, Annapia, Alessio Lorusso, Claudia Casaccia, Annamaria Conte, Federica Monaco, and Giovanni Savini. "Serum Neutralization Assay Can Efficiently Replace Plaque Reduction Neutralization Test for Detection and Quantitation of West Nile Virus Antibodies in Human and Animal Serum Samples." Clinical and Vaccine Immunology 21, no. 10 (August 6, 2014): 1460–62. http://dx.doi.org/10.1128/cvi.00426-14.

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ABSTRACTA serum neutralization assay (SN) was compared with the official plaque reduction neutralization test for the quantitation of West Nile virus antibodies. A total of 1,348 samples from equid sera and 38 from human sera were tested by these two methods. Statistically significant differences were not observed, thus supporting the use of SN for routine purposes.

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Zhang, Xi, and Binghua Ye. "Isolation of Prunin From Bauhinia variegata and Its Antioxidant Activity in Rats Fed an Atherogenic Diet." Natural Product Communications 15, no. 10 (October 2020): 1934578X2096787. http://dx.doi.org/10.1177/1934578x20967875.

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Atherosclerosis is no longer a disease attributed mainly to high cholesterol content in the body; it has come to be regarded as a chronic inflammatory disease with an autoimmune component. The purpose of this study was to investigate the effect of the prunin fraction (PF) isolated from the ethanolic extract of Bauhinia variegata against the release of various proinflammatory mediators in rats fed an atherogenic diet. The diet was administered orally to Sprague Dawley rats for 60 days to induce atherosclerosis. The blood serum of the rats was used to estimate the levels of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), thiobarbituric acid reactive substance, catalase, total cholesterol, triglyceride, low-density lipoprotein, and high-density lipoprotein using assay kits. Other physical parameters, such as body weight, feed intake, and systolic blood pressure, were also determined during the study. The results showed a significant protective effect of the PF against diet-induced atherosclerosis by decreasing the levels of proinflammatory mediators such as TNF-α and IL-6. Rats treated with PF (20 and 40 mg/kg) showed a change in systolic blood pressure and a reduction in oxidative stress induced by the atherogenic diet. Reduction in body weight and modulation of food intake were observed in PF-treated rats, which indicated atheroprotective, hypolipidemic, and antioxidant effects. The study concludes that the atheroprotective properties of PF are due to effects on the initial phase of plaque formation to thrombus formation. This study may help researchers to find a better alternative for selecting optimal therapies and preventing plaque formation. Future Significance: This article focuses on the molecular mechanisms involved in the evolution of atherosclerotic plaques and different targets that act at the starting stage of the plaque to thrombus formation. This may pave the way for selecting optimal therapies and preventing plaque complications.

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Abdel-Rahman, Adel A. H., Ibrahim F. Zeid, Hussien A. Barakat, and Wael A. El-Sayed. "Anti-Hepatitis B Virus Activity of New Substituted Pyrimidine Acyclic Nucleoside Analogues." Zeitschrift für Naturforschung C 64, no. 11-12 (December 1, 2009): 767–72. http://dx.doi.org/10.1515/znc-2009-11-1202.

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A number of N-substituted pyrimidine acyclic nucleosides were synthesized by coupling reaction of 2-(2-chloroethoxy)ethyl acetate or (2,2-dimethyl-1,3-dioxolan-4-yl)methyl 4-methylbenzenesulfonate with the corresponding base followed by deprotection. The synthesized compounds were tested for their antiviral activity against hepatitis B virus (HBV). The plaque reduction infectivity assay was used to determine virus count reduction as a result of treatment with the synthesized compounds which showed moderate to high antiviral activities

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Taylor, Shannon L., Paul R. Kinchington, Andrew Brooks, and Jennifer F. Moffat. "Roscovitine, a Cyclin-Dependent Kinase Inhibitor, Prevents Replication of Varicella-Zoster Virus." Journal of Virology 78, no. 6 (March 15, 2004): 2853–62. http://dx.doi.org/10.1128/jvi.78.6.2853-2862.2004.

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ABSTRACT Understanding the interactions between varicella-zoster virus (VZV) and host cells can be addressed by using small molecule inhibitors of cellular enzymes. Roscovitine (Rosco) is a purine derivative that inhibits cyclin-dependent kinase 1 (cdk1), cdk2, cdk5, cdk7, and cdk9, which are key regulators of the cell cycle and transcription. Herpesviruses are known to interact with cell cycle proteins; thus, the antiviral effects of Rosco on VZV growth were evaluated. In a plaque reduction assay, 25 μM Rosco prevented VZV replication, and the antiviral effect was reversible for at least up to 24 h posttreatment. Rosco also reduced expression of the major transactivator, IE62, over 48 h. Confocal microscopy studies indicated that Rosco caused the immediate-early proteins ORF4 and IE62 to abnormally localize in infected cells and prevented cell-cell spread of VZV over 48 h. Rosco was found to inhibit VZV DNA synthesis as measured by real-time PCR, and this technique was used to estimate the 50% effective concentration (EC50) of 14 μM. This value was close to the EC50 estimate of 12 μM determined from plaque reduction assays. At 25 μM, Rosco was not cytotoxic over 48 h in a neutral red uptake assay, and proliferation was slowed as the cells accumulated in a G2-like state. These results demonstrate the importance of cdk's in VZV replication and suggest that cdk inhibitors could serve as useful VZV antivirals.

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Hoshino, Y., M. M. Sereno, K. Midthun, J. Flores, R. M. Chanock, and A. Z. Kapikian. "Analysis by plaque reduction neutralization assay of intertypic rotaviruses suggests that gene reassortment occurs in vivo." Journal of Clinical Microbiology 25, no. 2 (1987): 290–94. http://dx.doi.org/10.1128/jcm.25.2.290-294.1987.

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Lee, Gunsup, Shailesh Budhathoki, Geum-Young Lee, Kwang-ji Oh, Yeon Ham, Young-Jun Kim, Ye Lim, et al. "Broad-Spectrum Antiviral Activity of 3D8, a Nucleic Acid-Hydrolyzing Single-Chain Variable Fragment (scFv), Targeting SARS-CoV-2 and Multiple Coronaviruses In Vitro." Viruses 13, no. 4 (April 9, 2021): 650. http://dx.doi.org/10.3390/v13040650.

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The virus behind the current pandemic, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for the etiology of novel coronavirus disease (COVID-19) and poses a critical public health threat worldwide. Effective therapeutics and vaccines against multiple coronaviruses remain unavailable. Single-chain variable fragment (scFv), a recombinant antibody, exhibits broad-spectrum antiviral activity against DNA and RNA viruses owing to its nucleic acid-hydrolyzing property. The antiviral activity of 3D8 scFv against SARS-CoV-2 and other coronaviruses was evaluated in Vero E6 cell cultures. Viral growth was quantified with quantitative RT-qPCR and plaque assay. The nucleic acid-hydrolyzing activity of 3D8 was assessed through abzyme assays of in vitro viral transcripts and cell viability was determined by MTT assay. We found that 3D8 inhibited the replication of SARS-CoV-2, human coronavirus OC43 (HCoV-OC43), and porcine epidemic diarrhea virus (PEDV). Our results revealed the prophylactic and therapeutic effects of 3D8 scFv against SARS-CoV-2 in Vero E6 cells. Immunoblot and plaque assays showed the reduction of coronavirus nucleoproteins and infectious particles, respectively, in 3D8 scFv-treated cells. These data demonstrate the broad-spectrum antiviral activity of 3D8 against SARS-CoV-2 and other coronaviruses. Thus, it could be considered a potential antiviral countermeasure against SARS-CoV-2 and zoonotic coronaviruses.

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Johnson, Alison J., Amanda J. Noga, Olga Kosoy, Robert S. Lanciotti, Alicia A. Johnson, and Brad J. Biggerstaff. "Duplex Microsphere-Based Immunoassay for Detection of Anti-West Nile Virus and Anti-St. Louis Encephalitis Virus Immunoglobulin M Antibodies." Clinical Diagnostic Laboratory Immunology 12, no. 5 (May 2005): 566–74. http://dx.doi.org/10.1128/cdli.12.5.566-574.2005.

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ABSTRACT West Nile (WN) virus was introduced into the United States in 1999, when the first human cases of WN fever and encephalitis appeared in New York City. From there, the virus has spread throughout North America, in some areas cocirculating with the related flavivirus St. Louis encephalitis (SLE) virus. Public health laboratories currently use an immunoglobulin M (IgM) antibody capture enzyme-linked immunosorbent assay (MAC-ELISA) as a primary test for human serodiagnosis, followed by a confirmatory plaque-reduction neutralization test (PRNT). The MAC-ELISAs take 2 days to perform; therefore there is a need for a more rapid test. This report describes a duplex microsphere-based immunoassay (MIA) that shortens the test processing time to about 4.5 h. The assay employs two sets of microspheres coupled to a single flavivirus group-reactive antibody, which are used to capture the WN and SLE viral antigens independently. Immunoglobulin G-depleted serum is concurrently assayed for IgM antibodies to each of the viral antigens. The results are standardized and classified by using quadratic discriminant analysis so that a single result, anti-WN IgM-positive, anti-SLE IgM-positive, negative, or nonspecific, can be determined. The duplex MIA results compared favorably to those of the plaque-reduction neutralization test and MAC-ELISA. The assay proved to be reproducible, produced accurate classifications as to the infecting virus, and was specific.

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Wang, Kuo Chih, Jung San Chang, Lien Chai Chiang, and Chun Ching Lin. "Cimicifuga foetida L. Inhibited Human Respiratory Syncytial Virus in HEp-2 and A549 Cell Lines." American Journal of Chinese Medicine 40, no. 01 (January 2012): 151–62. http://dx.doi.org/10.1142/s0192415x12500127.

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Human respiratory syncytial virus (HRSV) causes serious pediatric infection of the lower respiratory tract without effective therapeutic modality. Sheng-Ma-Ge-Gen-Tang (SMGGT; Shoma-kakkon-to) has been proven to be effective at inhibiting HRSV-induced plaque formation, and Cimicifuga foetida is the major constituent of SMGGT. We tested the hypothesis that C. foetida effectively inhibited the cytopathic effects of HRSV by a plaque reduction assay in both human upper (HEp2) and lower (A549) respiratory tract cell lines. Its ability to stimulate anti-viral cytokines was evaluated by an enzyme-linked immunosorbent assay (ELISA). C. foetida dose-dependently inhibited HRSV-induced plaque formation (p < 0.0001) before and after viral inoculation, especially in A549 cells (p < 0.0001). C. foetida dose-dependently inhibited viral attachment (p < 0.0001) and could increase heparins effect on viral attachment. In addition, C. foetida time-dependently and dose-dependently (p < 0.0001) inhibited HRSV internalization. C. foetida could stimulate epithelial cells to secrete IFN-β to counteract viral infection. However, C. foetida did not stimulate TNF-α secretion. Therefore, C. foetida could be useful in managing HRSV infection. This is the first evidence to support that C. foetida possesses antiviral activity.

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Sleeman, Katrina, Vasiliy P. Mishin, Varough M. Deyde, Yousuke Furuta, Alexander I. Klimov, and Larisa V. Gubareva. "In Vitro Antiviral Activity of Favipiravir (T-705) against Drug-Resistant Influenza and 2009 A(H1N1) Viruses." Antimicrobial Agents and Chemotherapy 54, no. 6 (March 29, 2010): 2517–24. http://dx.doi.org/10.1128/aac.01739-09.

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ABSTRACT Favipiravir (T-705) has previously been shown to have a potent antiviral effect against influenza virus and some other RNA viruses in both cell culture and in animal models. Currently, favipiravir is undergoing clinical evaluation for the treatment of influenza A and B virus infections. In this study, favipiravir was evaluated in vitro for its ability to inhibit the replication of a representative panel of seasonal influenza viruses, the 2009 A(H1N1) strains, and animal viruses with pandemic (pdm) potential (swine triple reassortants, H2N2, H4N2, avian H7N2, and avian H5N1), including viruses which are resistant to the currently licensed anti-influenza drugs. All viruses were tested in a plaque reduction assay with MDCK cells, and a subset was also tested in both yield reduction and focus inhibition (FI) assays. For the majority of viruses tested, favipiravir significantly inhibited plaque formation at 3.2 μM (0.5 μg/ml) (50% effective concentrations [EC50s] of 0.19 to 22.48 μM and 0.03 to 3.53 μg/ml), and for all viruses, with the exception of a single dually resistant 2009 A(H1N1) virus, complete inhibition of plaque formation was seen at 3.2 μM (0.5 μg/ml). Due to the 2009 pandemic and increased drug resistance in circulating seasonal influenza viruses, there is an urgent need for new drugs which target influenza. This study demonstrates that favipiravir inhibits in vitro replication of a wide range of influenza viruses, including those resistant to currently available drugs.

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Gibbs, Samantha E. J., Douglas M. Hoffman, Lillian M. Stark, Nicole L. Marlenee, Bradley J. Blitvich, Barry J. Beaty, and David E. Stallknecht. "Persistence of Antibodies to West Nile Virus in Naturally Infected Rock Pigeons (Columba livia)." Clinical Diagnostic Laboratory Immunology 12, no. 5 (May 2005): 665–67. http://dx.doi.org/10.1128/cdli.12.5.665-667.2005.

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ABSTRACT Wild caught rock pigeons (Columba livia) with antibodies to West Nile virus were monitored for 15 months to determine antibody persistence and compare results of three serologic techniques. Antibodies persisted for the entire study as detected by epitope-blocking enzyme-linked immunosorbent assay and plaque reduction neutralization test. Maternal antibodies in squabs derived from seropositive birds persisted for an average of 27 days.

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Liu, Lidong, Kun Wen, Jie Li, Dongmei Hu, Yanfen Huang, Liwen Qiu, Jianpiao Cai, and Xiaoyan Che. "Comparison of Plaque- and Enzyme-Linked Immunospot-Based Assays To Measure the Neutralizing Activities of Monoclonal Antibodies Specific to Domain III of Dengue Virus Envelope Protein." Clinical and Vaccine Immunology 19, no. 1 (November 23, 2011): 73–78. http://dx.doi.org/10.1128/cvi.05388-11.

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ABSTRACTThe plaque reduction neutralization test (PRNT) is used widely to measure the neutralization activity of anti-dengue virus (DENV) antibodies, but it is time-consuming and labor-intensive and has low sample throughput. For fast and convenient measurement of neutralizing antibodies, especially in evaluating the efficiency of the DENV vaccines on a large scale, a new method is needed to replace PRNT. In recent decades, several microneutralization assays have been developed to overcome the limitations of PRNT. In the present study, we evaluated one of these, the enzyme-linked immunospot microneutralization test (ELISPOT-MNT), in comparison with PRNT. ELISPOT-MNT is performed in 96-well format, and the plaques are developed after 2 to 4 days using an ELISA to transform them into spots, which are detected automatically with an ELISPOT instrument. The assay is faster than PRNT, has a high throughput, and is more objective. We used 10 monoclonal antibodies (MAbs) against domain III of the DENV envelope protein (EDIII) to evaluate the two assays; all of these MAbs cross-react with all four serotypes of DENV as measured by immunofluorescence assay. The two neutralization assays were performed simultaneously to measure the 50% inhibitory concentration (IC50) of these MAbs. Using PRNT as the reference and treating IC50values higher than 50 μg/ml of MAbs as negative, ELISPOT-MNT showed a sensitivity of 95.6% and specificity of 88.24% when 10 MAbs were tested against four DENV serotype strains. A good correlation (R2= 0.672;P= 0.000) was observed between the two assays, making ELISPOT-MNT a potentially valuable method for measure of neutralizing antibodies against DENV.

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Vicenti, Ilaria, Filippo Dragoni, Alessia Giannini, Federica Giammarino, Michele Spinicci, Francesco Saladini, Adele Boccuto, and Maurizio Zazzi. "Development of a Cell-Based Immunodetection Assay for Simultaneous Screening of Antiviral Compounds Inhibiting Zika and Dengue Virus Replication." SLAS DISCOVERY: Advancing the Science of Drug Discovery 25, no. 5 (March 18, 2020): 506–14. http://dx.doi.org/10.1177/2472555220911456.

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Practical cell-based assays can accelerate anti-Zika (ZIKV) and anti-dengue (DENV) virus drug discovery. We developed an immunodetection assay (IA), using a pan-flaviviral monoclonal antibody recognizing a conserved envelope domain. The final protocol includes a direct virus yield reduction assay (YRA) carried out in the human Huh7 cell line, followed by transfer of the supernatant to a secondary Huh7 culture to characterize late antiviral effects. Sofosbuvir and ribavirin were used to validate the assay, while celgosivir was used to evaluate the ability to discriminate between early and late antiviral activity. In the direct YRA, at 100, 50, and 25 TCID50, sofosbuvir IC50 values were 5.0 ± 1.5, 2.7 ± 0.5, 2.5 ± 1.1 µM against ZIKV and 16.6 ± 2.8, 4.6 ± 1.4, 2.6 ± 2.2 µM against DENV; ribavirin IC50 values were 6.8 ± 4.0, 3.8 ± 0.6, 4.5 ± 1.4 µM against ZIKV and 17.3 ± 4.6, 7.6 ± 1.2, 4.1 ± 2.3 µM against DENV. Sofosbuvir and ribavirin IC50 values determined in the secondary YRA were reproducible and comparable with those obtained by direct YRA and plaque reduction assay (PRA). In agreement with the proposed mechanism of late action, celgosivir was active against DENV only in the secondary YRA (IC50 11.0 ± 1.0 µM) and in PRA (IC50 10.1 ± 1.1 µM). The assay format overcomes relevant limitations of the gold standard PRA, allowing concurrent analysis of candidate antiviral compounds against different viruses and providing preliminary information about early versus late antiviral activity.

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Journal articles on the topic 'Plaque reduction assay'

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