scholarly journals Detection of Polioviruses in Sewage Using Cell Culture and Molecular Methods

2026 ◽  
Vol 65 (4) ◽  
pp. 479-483
Author(s):  
Agnieszka Figas ◽  
Magdalena Wieczorek ◽  
Bogumiła Litwińska ◽  
Włodzimierz Gut
Keyword(s):  
Cell Culture ◽  
Cell Lines ◽  
Cell Cultures ◽  
Environmental Samples ◽  
Genetic Material ◽  
Culture Technique ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Step Algorithm ◽  
Cell Culture Technique

The work presented here demonstrates the utility of a two-step algorithm for environmental poliovirus surveillance based on: preselection of sewage samples tested for the presence of enteroviral genetic material-RT-PCR assay and detection of infectious viruses by cell culture technique (L20B for polioviruses and RD for polio and other non-polio enteroviruses). RD and L20B cell lines were tested to determine their sensitivity for isolation of viruses from environmental samples (sewage). Finally, we wanted to determine if sewage concentration affects the results obtained for RT-PCR and cell cultures.

scholarly journals Detection of Infectious Adenovirus in Cell Culture by mRNA Reverse Transcription-PCR

2003 ◽  
Vol 69 (12) ◽  
pp. 7377-7384 ◽  
Author(s):  
Gwangpyo Ko ◽  
Theresa L. Cromeans ◽  
Mark D. Sobsey
Keyword(s):  
Cell Culture ◽  
Cell Cultures ◽  
Reverse Transcription ◽  
Radiation Detection ◽  
Virus Infectivity ◽  
Mrna Transcript ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Mrna Detection ◽  
Primer Sets

ABSTRACT We have developed and evaluated the reverse transcription (RT)-PCR detection of mRNA in cell culture to assay infectious adenoviruses (Ads) by using Ad type 2 (Ad2) and Ad41 as models. Only infectious Ads are detected because they are the only ones able to produce mRNA during replication in cell culture. Three primer sets for RT-PCR amplification of mRNA were evaluated for their sensitivity and specificity: a conserved region of late mRNA transcript encoding a virion structural hexon protein and detecting a wide range of human Ads and two primer sets targeting a region of an early mRNA transcript that specifically detects either Ad2 and Ad5 or Ad40 and Ad41. The mRNAs of infected A549 and Graham 293 cells were recovered from cell lysates with oligo(dT) at different time periods after infection and treated with RNase-free DNase to remove residual contaminating DNA, and then Ad mRNA was detected by RT-PCR assay. The mRNA of Ad2 was detected as early as 6 h after infection at 106 infectious units (IU) per cell culture and after longer incubation times at levels as low as 1 to 2 IU per cell culture. The mRNA of Ad41 was detected as soon as 24 h after infection at 106 IU per cell culture and at levels as low as 5 IU per cell culture after longer incubation times. To confirm the detection of only infectious viruses, it was shown that no mRNA was detected from Ad2 and Ad41 inactivated by free chlorine or high doses of collimated, monochromatic (254-nm) UV radiation. Detection of Ad2 mRNA exactly coincided with the presence of virus infectivity detected by cytopathogenic effects in cell cultures, but mRNA detection occurred sooner. These results suggest that mRNA detection by RT-PCR assay in inoculated cell cultures is a very sensitive, specific, and rapid method by which to detect infectious Ads in water and other environmental samples.

Polymerase chain reaction amplification and typing of rotavirus in environmental samples

1995 ◽  
Vol 31 (5-6) ◽  
pp. 371-374 ◽  
Author(s):  
R. Gajardo ◽  
R. M. Pintó ◽  
A. Bosch
Keyword(s):  
Polymerase Chain Reaction ◽  
Environmental Samples ◽  
Polymerase Chain Reaction Amplification ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Pcr Assay ◽  
Group A ◽  
Reaction Amplification ◽  
Stool Specimens ◽  
Polymerase Chain

A reverse transcription polymerase chain reaction (RT-PCR) assay is described that has been developed for the detection and serotyping of group A rotavirus in stool specimens and concentrated and non-concentrated sewage specimens.

A Cell Culture Technique for Chromosome Preparation in Cyprinid Fishes

Copeia ◽  
1984 ◽  
Vol 1984 (1) ◽  
pp. 232 ◽  
Author(s):  
Chris T. Amemiya ◽  
John W. Bickham ◽  
John R. Gold
Keyword(s):  
Cell Culture ◽  
Culture Technique ◽  
Chromosome Preparation ◽  
A Cell ◽  
Cyprinid Fishes ◽  
Cell Culture Technique

Antimigratory effect of berberine in T98G, U87MG and primary glioma cell culture.

2021 ◽  
Vol 39 (15_suppl) ◽  
pp. e15045-e15045
Author(s):  
Irina V. Mezhevova ◽  
Svetlana Yu. Filippova ◽  
Sofia V. Timofeeva ◽  
Anastasia O. Sitkovskaya ◽  
Tatiana V. Shamova ◽  
...  
Keyword(s):  
Cell Culture ◽  
Cell Lines ◽  
Cell Cultures ◽  
Glioma Cell ◽  
Glioma Cells ◽  
Surgical Removal ◽  
Experimental Sample ◽  
Wound Area ◽  
Migratory Activity ◽  
Primary Glioma

e15045 Background: Berberine is an alkaloid compound with a structure that is highly similar to that of intercalating agents. It affects numerous cell signaling pathways and is widely studied as potential anticancer drug. It is known that berberine affects cancer cells migration through metalloproteinase-2 inhibition, but this effect was never studied on glioma cells. Anti-migratory drugs are of special interest in brain cancer therapy since glioma's highly invasive nature makes total surgical removal of tumor practically impossible. The aim of the study was to evaluate berberine anti-migratory activity on glioma cells. Methods: Cell migration capacity of T98G and U87MG cell lines, as well as primary glioma cell culture established in our laboratory, was assessed via standard wound healing assay with automated image acquisition and analysis on Lionheart FX (BioTek) cell imager. Prior to assay setting up cell cultures were maintained in DMEM medium with L-glutamine (1 μM) (Gibco) and 10% FBS (Gibco) at 37C0 and 5.0% CO2. Cells were seeded at 250 000 cells per well on 24-well plates and incubated overnight in order to attach to plate bottom. After that a vertical wound was made manually in each well, and berberine was added to experimental wells to final concentration 50 mg/L. Plates with cells were continuously incubated and photographed in cell imager at 37C0 and 5.0% CO2. The extent of cells migration was measured as the percent of wound area decrease after 24 hours of incubation in relation to starting time point. Data are given as: Mean ± 95% confidence interval. Results: In our study we berberine exhibited anti-migratory activity in all cell cultures under study. In rather fast growing primary cell culture wound area decrease was 99.23%±0.62% in control sample and 91.75%±0.28% in experimental sample. The difference was small but significant at p < 0.001 level (df = 30). Popular permanent glioma cell lines T98G and U87MG showed more prominent decrease in studied parameter with higher degree of variance at the same time. In T98G wound area decrease was 71.6%±12.3% in control and 48.8%± 7.6% in experimental samples after 24 hours of cultivation in presence of 50 mg/L berberine. While U87MG demonstrated 60.28%±5.13% and 37.5%± 8.34% wound area decrease accordingly. The obtained difference between control and experimental groups in permanent cell cultures was statistically significant at the 0.05 level (df = 30). Conclusions: Our preliminary research proved berberine to be potent anti-migratory agent in glioma treatment. Further investigations are needed to evaluate its ability to inhibit glioma cell expansion in vivo.

scholarly journals The Detection of Enteroviruses in Sewage Using Caco-2 Cells

2013 ◽  
Vol 62 (1) ◽  
pp. 97-100 ◽  
Author(s):  
MAGDALENA WIECZOREK ◽  
ŁUKASZ KURYK ◽  
AGNIESZKA WITEK ◽  
ANNA DIUWE ◽  
BOGUMIŁA LITWIŃSKA
Keyword(s):  
Cell Culture ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Beef Extract

The work presented here demonstrates the utility of Caco-2 cells to detect enteroviruses in sewage. Viruses were concentrated by beef extract elution and organic flocculation prior to analysis by cell culture assays and RT-PCR. Enteroviruses were detected in all sewage samples, but only one sample was positive solely in RT-PCR assay. We proved that Caco-2 cells were more effective than RD and L20B cells in enterovirus isolation, depending on procedures used in the inoculation process.

scholarly journals Virological applications of the grid-cell-culture technique

1990 ◽  
Vol 3 (1) ◽  
pp. 1-27 ◽  
Author(s):  
Alex D. Hyatt ◽  
Bryan T. Eaton
Keyword(s):  
Cell Culture ◽  
Grid Cell ◽  
Culture Technique ◽  
Cell Culture Technique

Development of an integrated cell culture—Real-time RT-PCR assay for detection of reovirus in biosolids

2007 ◽  
Vol 139 (2) ◽  
pp. 195-202 ◽  
Author(s):  
Elizabeth M. Gallagher ◽  
Aaron B. Margolin
Keyword(s):  
Cell Culture ◽  
Real Time ◽  
Rt Pcr ◽  
Pcr Assay

scholarly journals Evaluation of the potential role of long non-coding RNA LINC00961 in luminal breast cancer: a case–control and systems biology study

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Sepideh Mehrpour Layeghi ◽  
Maedeh Arabpour ◽  
Rezvan Esmaeili ◽  
Mohammad Mehdi Naghizadeh ◽  
Javad Tavakkoly Bazzaz ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Cell Lines ◽  
Potential Role ◽  
Bioinformatics Analysis ◽  
T Test ◽  
Rt Pcr ◽  
Pcr Assay ◽  
Luminal A ◽  
Ppi Networks

Abstract Background Luminal subtype is the most common subgroup of breast cancer (BC), accounting for more than 70% of this cancer. Long non-coding RNAs (lncRNAs) are a group of RNAs which play critical roles in diverse cellular processes. It is proved that dysregulation of them can contribute to the development of various cancers, including BC. LINC00961 was reported to be downregulated in several cancers, however, its expression level in BC remains largely unknown. The purpose of the present study was to investigate the possible role of LINC00961 in luminal A and B subtypes of BC. Methods To obtain novel lncRNAs associated with different cancers and differentially expressed lncRNAs (DElncRNAs) between BC tumor and normal tissues, Lnc2Cancer and GDC databases were used, respectively. After performing literature review, the expression level of the selected lncRNA (LINC00961) was evaluated in 79 luminal A and B BC specimens and adjacent non-cancerous tissues by Quantitative Reverse Transcription PCR (qRT-PCR). LINC00961 expression was also evaluated in two luminal A BC cell lines, compared to a normal breast cell line. The comparison of the differences between tumor and adjacent non-tumor samples was performed by paired sample t-test. Moreover, correlation analysis between LINC00961 expression and clinicopathological features was performed using the chi-square, fisher exact, and independent t-test. In order to investigate the possible roles of LINC00961 in luminal A and B BC, different bioinformatics analyses such as functional annotation of the LINC00961 co-expressed genes and protein–protein interaction (PPI) networks construction were also performed. Results LINC00961 was selected as a significant DElncRNA which had not been studied in BC. According to q-RT PCR assay, LINC00961 was downregulated in luminal BC tissues and cell lines. Its expression was correlated with smoking status and the age of menarche in luminal BC patients. Also, the results of the bioinformatics analysis were consistent with the data obtained from q-RT PCR assay. The final results indicated that LINC00961 might be involved in multiple cancer-associated pathways such as chemokine, Ras and PI3K–Akt signaling pathways, GPCR ligand binding, and signal transduction in luminal subtypes of BC. CDH5, GNG11, GNG8, SELL, S1PR1, CCL19, FYN, ACAN, CD3E, ACVRL1, CAV1, and PPARGC1A were identified as the top hub genes of the PPI networks across luminal subgroup. Conclusion Our findings suggested that LINC00961 was significantly downregulated in luminal A and B subtypes of BC. Moreover, bioinformatics analysis provided a basis for better identification of the potential role of LINC00961 in luminal subtype of BC.

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