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RISS 인기검색어
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- 저자 : 손여원(Yeowon Sohn) (검색결과 2 건)
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외래성 Minute Virus of Mice 검출을 위한 시험법 개발에 관한 연구
정지원(Jeewon Joung),이숙연(Sookyeon Lee),장석기(Seokkee Chang),정자영(Jayoung Jeong),손여원(Yeowon Sohn) 한국실험동물학회 2005 Laboratory Animal Research Vol.21 No.1
In manufacturing of biotechnological products, numerous adventitious agents might have been detected in cell substrates. Recently a biotech-pharmaceutical company has had experienced with contamination of large-scale cell cultures by minute virus of mice (MVM) and this resulted in both the loss of product and the need for major cleaning validation procedures to be put in place. We have developed a simple PCR assay to detect the presense of MVM in cell culture supernatant. The specificity of MVM was determined by testing viral preparations of other autonomous parvovirus and MVM specific PCR assay detects only in the presence MVM viral DNA. This assay has proven to be very sensitive and it can detect 10 or fewer genome equivalents (copies) of MVM. These results implicate that we may use this assay for routine screening test for MVM. The assay also can detect MVM following extraction procedure of DNA in cell lines at a concentration of 1 TCID₅₀/㎖ and establishes a sensitive, specific assay that can detect the presense of MVM sequences in a cell culture system.
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HIV-1 유래 렌티바이러스 벡터의 복제가능 바이러스 검출과 역가측정 분석방법 비교
장석기(Seok Kee Chang),오일웅(Il Ung Oh),정자영(Jayoung Jeong),안광수(Kwang Soo Ahn),손여원(Yeowon Sohn) 대한약학회 2005 약학회지 Vol.49 No.3
Human Immunodeficiency Virus type 1 (HIV-1) based lentivirus vector has demonstrated great potential as gene therapy vectors mediating efficient gene delivery and long-term transgene expression in both dividing and nondividing cells. However, for clinical studies it must be confirmed that vector preparations are safe and not contaminated by replication competent lentivirus (RCL) related to the parental pathogenic virus, HIV-1. In this study, we would like to establish the method for titration and RCL detection of lentivirus vector. The titration was determined by vector expression containing the green fluorescent protein, GFP in transduced cells. The titer was 1×107 Transducing Unit/ml in the GFP expression assay and 8.9×107 molecules/ml in the real-time PCR. Also, for the detection of RCL, we have used a combination method of PCR and p24 antigen detection. First, PBS/psi and VSV-G region in the genomic DNA of transduced cells was detected by PCR assay. Second, transfer and expression of the HIV-1 gag gene was detected by p24 ELISA. In an attempt to amplify any RCL, the transduced cells were cultured for 3 weeks (amplification phase) and the supernatant of amplified transduced cell was used for the second transduction to determine whether a true RCL was present (indicator phase). Analysis of cells and supernatant at day 6 in indicator phase were negative for PBS/psi, VSV-G, and p24 antigen. These results suggest that they are not mobilized and therefore there are no RCL in amplification phase. Thus, real-time PCR is a reliable and sensitive method for titration and RCL detection of lentivirus vector.
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