• Korean Journal of Veterinary Service
• /
• 제45권1호
• /
• pp.19-30
• /
• 2022

In this study, a simple loop-mediated isothermal amplification (LAMP) combined with visual detection method (vLAMP) assay was developed for the rapid and specific detection of African swine fever virus (ASFV), overcoming the shortcomings of previously described LAMP assays that require additional detection steps or pose a cross-contamination risk. The assay results can be directly detected by the naked eye using hydroxynaphthol blue after incubation for 40 min at 62℃. The assay specifically amplified ASFV DNA and no other viral nucleic acids. The limit of detection of the assay was <50 DNA copies/reaction, which was ten times more sensitive than conventional polymerase chain reaction (cPCR) and comparable to real-time PCR (qPCR). For clinical evaluation, the ASFV detection rate of vLAMP was higher than cPCR and comparable to OIE-recommended qPCR, showing 100% concordance, with a κ value (95% confidence interval) of 1 (1.00~1.00). Considering the advantages of high sensitivity and specificity, no possibility for cross-contamination, and being able to be used as low-cost equipment, the developed vLAMP assay will be a valuable tool for detecting ASFV from clinical samples, even in resource-limited laboratories.

• Korean Journal of Veterinary Research
• /
• 제36권2호
• /
• pp.277-281
• /
• 1996

A technique to improve the analysis of micronuclei(MN) in lymphocytes as a cytogenetic indicator is reported. For the purpose of diminishing the variation of the result from individual reader and making it easier to distinguish accurately a cytokinesis blicked(CB) lymphocyte and micronuclei, we tried a modified one-step silver staining technique as a method for detection of the argyrophilic nucleolar organizer region(AgNOR) with or without conventional Giemsa stain in the slide from CB method. Compared with the conventional Giemsa stain, the preparation processed with this method are especially useful for the accurate analysis of MN of cultured lymphocyte with cytochalasin B. This method will be a useful technique for automated calculation of MN.

• Clinical and Experimental Reproductive Medicine
• /
• 제16권2호
• /
• pp.173-177
• /
• 1989

Defective or inadequate semen quality, usually presenting as low sperm count or poor sperm motility , is recognizable by semen analysis. However, the ability of spermatozoa to fertilize an ovum is not determined used in various experiments. In this study, hamster oocyte sperm penetration assay was used to determine the fertilizing capacity of sperms in 20 subjects which divided into two groups, group A with 10 normal fertile men, and group B with 10 infertile men. The % penetration in group A and group B were 61% and 35% respectively, which showed statistically not significant but fertilization index was significantly different between group A(FI=2.24) and group B(FI=O.05). Additionally it seemed that the percentage of sperm penetraton was influenced more by the motility of spermatozoa than by the number.

• Proceedings of the PSK Conference
• /
• 대한약학회 2002년도 Proceedings of the Convention of the Pharmaceutical Society of Korea Vol.2
• /
• pp.406.2-406.2
• /
• 2002

Special attention has been paid to human chorionic gonadotropin (hCG) for athlete doping control because it stimulates the endogenous production of testosterone and epitestosterone without increasing the T/E ratio which is a doping indicator for the exogenous administration of testosterone. Even though the IOC banned the use of hCG. a detection method has not been decided upon since there are a variety of immunoassay kits available on the market. We evaluated three kits in terms af their performance characteristics. (omitted)

• Journal of Nutrition and Health
• /
• 제29권9호
• /
• pp.971-980
• /
• 1996

The soluble transferrin receptor(TfR) in human serum has been shown recently to be a truncated form of intact membrane bound receptor containing most of the extracellular domain. We purfied the transferin-free TfR from human serum by immounoaffinity chromatography which produced the single protein identity in high resolution gel chormatography. The monoclonal antibodies(MAb) against purifed serum TfR were produced by fusion of spleen cells o fimmunized Balb/c mice and SP2 cells. Ten hybrids producing MAb specific for serum TfR were identifed and determine their iostypes. A immunoraddiometric assay (IRMA) for serum TfR was established using two monoclonal IgG1 antibodies as the coating and indicator antibodies on the bosis of their suitability in sandwich IRMA of serum TfR. The mean serum TfR levels in the 15 normal male, 15 normal female, and 19 iron-deficient subjects were 5.4$\pm$0.98, 4.6$\pm$0/76, and 18.0$\pm$12.8mg/1, respectively, and the difference in mean values between normal and iron deficient subjects was significant(p=0.0005). There existed the inverse logarithmic relationship(r=-0.9336, p<0.0001) between the serum TfR and ferritin levels.

• The Journal of the Korean Society for Microbiology
• /
• 제22권2호
• /
• pp.163-166
• /
• 1987

A new method for rosette assay is described for the detection of antigen-specific lymphocytes from BCG-infected mice using sheep erythrocytes coated with BCG antigen. The optimal concentration of BCG antigen for preparation of indicator cells and the incubation time of antigen coated erythrocytes-lymphocytes mixture were $50\;{\mu}g/ml$ and 1 h, respectively. The number of rosette-forming cells (RFC) during the course of BCG infection showed gradual increase as infection progressed and RFC was reached maximum (about 5-7% of splenic lymphocytes formed rosette) at 3 or 4 weeks after infection.

• Journal of Microbiology and Biotechnology
• /
• 제29권10호
• /
• pp.1543-1552
• /
• 2019

Salmonella is a common zoonotic and foodborne pathogen that causes high morbidity and mortality in developing countries. In this study, we established and validated a polymerase spiral reaction (PSR) assay which targeted the conserved invasion gene (invA) of Salmonella by SYBR Green I indicator methods. Subsequently, assays for determination of the optimal conditions for optimal specificity and sensitivity of PSR were performed. We performed comprehensive evaluations using loop-mediated isothermal amplification (LAMP) and real-time PCR. A total number of 532 samples of daily food were analyzed by PSR. Twenty-seven bacterial strains were tested in the specificity assay, from which positive results were obtained only for 14-Salmonella strains. However, none of the 13 non-Salmonella strains was amplified. Similarly with LAMP and real-time PCR, the detection limit of the PSR assay was 50 CFU/ml. The PSR method was also successfully applied to evaluate the contamination with Salmonella in 532 samples of daily food, corroborating traditional culture method data. The novel PSR method is simple, sensitive, and rapid and provides new insights into the prevention and detection of foodborne diseases.

• YAKHAK HOEJI
• /
• 제49권3호
• /
• pp.217-224
• /
• 2005

Human Immunodeficiency Virus type 1 (HIV-l) 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-l. 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 $1{\times}10^7$ Transducing Unit/ml in the GFP expression assay and $8.9{\times}10^7$ 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.

• Korean Journal of Clinical Laboratory Science
• /
• 제41권1호
• /
• pp.37-41
• /
• 2009

Measurement of hemoglobin A1c is used as an objective indicator of long-term blood glucose control in diabetic patients. We evaluated recently introduced Norudia$^{(R)}$ HbA1c (Daiichi Pure Chemical Co. Ltd, Tokyo, Japan) test reagent using enzyme method for HbA1c assay. Linearity, precision and correlation with VARIANT$^{TM}$ II Turbo HbA1c analyzer (BIO-RAD, Hercules, CA, USA) were evaluated. The reference range was determined from 201 healthy subjects. The Norudia$^{(R)}$ HbA1c test reagent was founded to be linear in a range of 5.6% to 14.0% ($r^2=0.9885$). The within-run and between-day precision were 0.954% and 1.03% for low level (HbA1c 5.24%), 0.67% and 1.28% for high level (HbA1c 9.01%), respectively. Comparison study between Norudia$^{(R)}$ HbA1c test reagent and VARIANT$^{TM}$ II Turbo showed good correlation with a slope of 1.0489. an intercept at -0.9717, and coefficient of correlation was 0.9907. The reference range of HbA1c obtained from this reagent was 4.07-5.50%. The Norudia$^{(R)}$ HbA1c test reagent showed good linearity, precision and correlation with HbA1c analyzer with HPLC method. In addition, the exclusive analyzer is not required for assay and then this kit may be useful for HbA1c assay in clinical laboratory.

• Biomolecules & Therapeutics
• /
• 제10권3호
• /
• pp.186-192
• /
• 2002

Special attention has been paid to human chorionic gonadotropin (hCG) for athlete doping control because it stimulates the endogenous production of testosterone and epitestosterone without increasing the T/E ratio which is a doping indicator for the exogenous administration of testosterone. Even though the IOC banned the use of hCG, a detection method has not been decided upon since there are a variety of immunoassay kits available on the market. We evaluated three kits in terms of their performance characteristics. The assay value of the control sample varied depending on the kit, resulting in 198 mIU/ml for the MAIA kit, 172mIU/ml for the IRMA kit, and 143 mIU/ml for the MEIA kit. Considering the IOC inter-lab distribution of results(55-312 mIU/ml) using 27 different kits and the mean value (178$\pm$56 mIU/ml), all three kits are within the range of －15.8% - ＋5.6% of the mean value, which proves them useful for the hCG assay. The MEIA kit resulted in lower hCG values because it detects only intact hCG molecules, in contrast to the other two kits which detect intact hCG and -hCG together. However, it is suitable for screening purposes because of its advantage of being an automated system. When 123 urine samples of athletes were analyzed in 22 batches using this system, the variation of control values fell within $\pm$ 10% of the mean values, and an specimens tested negative with hCG values less than the detection limit of 2 mIU/ml.