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논문 기본 정보
- 자료유형
- 학술저널
- 저자정보
- 발행연도
- 2017.10
- 수록면
- 307 - 316 (10page)
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초록· 키워드
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Quantitative PCR and plaque assay are powerful virological techniques used to measure the load of defective or infectious virus in mouse and human. However, these methods display limitations such as cross contamination and long run-time. Here, we describe a novel technique termed as semi-functional quantitative flow cytometry (SFQF) for the accurate estimation of the quantity of infectious lymphocytic choriomeningitis virus (LCMV). LCMV titration method using flow cytometry was previously developed but has technical shortcomings, owing to the less optimized parameters such as cell overgrowth, plate scale, and detection threshold. Therefore, we first established optimized conditions for SFQF assay using LCMV nucleoprotein (NP)-specific antibody to evaluate the threshold of the virus detection range in the plaque assay. We subsequently demonstrated that the optimization of the method increased the sensitivity of virus detection. We revealed several new advantages of SFQF assay, which overcomes some of the previously contentious points, and established an upgraded version of the previously reported flow cytometric titration assay. This method extends the detection scale to the level of single cell, allowing extension of its application for in vivo detection of infected cells and their phenotypic analysis. Thus, SFQF assay may serve as an alternative analytical tool for ensuring the reliability of LCMV titration and can be used with other types of viruses using target-specific antibodies.
목차
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS
DISCUSSION
REFERENCES
참고문헌 (24)
참고문헌 신청
Zhou X, Ramachandran S, Mann M, Popkin DL. Role of lymphocytic choriomeningitis virus (LCMV) in understanding viral immunology: past, present and future. Viruses 2012;4:2650-2669.
Salvato MS, Shimomaye EM. The completed sequence of lymphocytic choriomeningitis virus reveals a unique RNA structure and a gene for a zinc finger protein. Virology 1989;173:1-10.
Pinschewer DD, Perez M, de la Torre JC. Role of the virus nucleoprotein in the regulation of lymphocytic choriomeningitis virus transcription and RNA replication. J Virol 2003;77:3882-3887.
Darbre S, Johnson S, Kallert S, Lambert PH, Siegrist CA, Pinschewer DD. The nucleoprotein is required for lymphocytic choriomeningitis virus-based vaccine vector immunogenicity. J Virol 2015;89:11734-11738.
Marcus PI, Carver DH. Hemadsorption-negative plaque test: new assay for rubella virus revealing a unique interference. Science 1965;149:983-986.
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UCI(KEPA) : I410-ECN-0101-2018-517-001400672