• Title/Summary/Keyword: Enzyme Linked Immunosorbent Assay(ELISA)

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ELISA of paragonimiasis in cat by crude and purified antigens of Paragonimus westermani (폐흡충(肺吸蟲)의 조(粗)항원과 정제(精製)항원에 의한 폐흡충(肺吸蟲)감염 고양이혈청의 면역효소반응(免疫酵素反應))

  • Lee, Ok-Ran;Chang, Jae-Kyung
    • The Korean Journal of Parasitology
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    • v.24 no.2
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    • pp.187-193
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    • 1986
  • Enzyme-linked immunosorbent assay(ELISA) using crude and affinity-purified antigens of adult worms of Paragonimus westermani was performed for infected cat sera with different worm burden, from preinfection to 18th week after infection. Crude antigen was used with supernatant of homogenated worms by freezing-thawing method, and the supernate was centrifuged for 1 hour at 10,000 rpm at $4^{\circ}C$. Affinity-purified antigen(antibody-bound antigen) was prepared from fractions(bound and unbound) of crude antigen by affinity chromatography on CNBr-activated sepharose 4B, and IgG as a ligand was prepared from paragonimiasis cat serum(6 months infected) obtained by ammonium sulfate ($40%{\sim}45%$ saturated) precipitation method. By SDS-PAGE, crude antigen showed 22 polypeptide fractions while purified antigen showed 4 fractions: 36, 400, 34, 700, 27, 600 and 11, 500 in molecular weights. All cats were divided into five groups($G_1-G_5$) by different worm burdens. The mean of recovered worms(${\pm}SD$) and the number of cats in each group are as follows: $G_1$, 2 worms(0) and 4 cats; $G_2$, 4.75 (${\pm}0.66$) and eight; $G_3$, 10.75(${\pm}1.92$) and four; $G_4$, 23.20(${\pm}3.43$) and five; $G_5$, 48(${\pm}12.63$) and five cats. The results were summarized as follows: 1. The antibody levels(OD value) increased by worm burden in $G_1$ to $G_4$ generally. However, individual antibody levels were not exactly related with worm burden in all groups, especially there was a wide difference in $G_4$ and $G_5$. These results suggested that the worm burden in $G_4$ (about $20{\sim}30$ worms) is enough to produce antibody maximum in cats of $2{\sim}3kg$ weight. 2. The antibody levels increased significantly(p<0.05) compared to control sera at the 3rd week in $G_1$ and $G_2$, at the 2nd week in $G_3$, and at the 1st week in $G_4$ and $G_5$. Especially in the 4th week, OD value increased more in $G_1$(p<0.01) and in $G_2$ to $G_5$(p<0.001). In the pattern of antibody levels by ELISA in each group, OD in $G_1$ increased to the 18th week continuously, in $G_2$ OD was maintained same after the 16th week, but in $G_3$ it decreased after the 16th week, and it was maintained same in $G_4$ and $G_5$ after the 14th week. 3. The antibody levels by ELISA with the affinity-purified antigen were higher than those with crude antigen in all groups generally. Especially, the difference of OD values between two antigens was larger from the 4th to the 10th week. In $G_1$ and $G_2$ OD with purified antigen was higher than that with crude one to the 18th week. It was also higher in $G_3$ than that with crude antigen to the 16th week and OD of $G_4$ and $G_5$ were higher before the 14th week than that with crude antigen, however became lower at the 16th week. Consequently, the antibody level in ELISA with affinity-purified antigen was more sensitive at the early weeks after infection and in light infection groups than that with crude antigen.

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Can Peritoneal Dialysis Remove Endothelin-1 after Cardiopulmonary Bypass for Repair of Congenital Heart Disease? (체외순환을 이용한 선천성 심장수술 후에 시행한 복막투석이 내피의존성 혈관 활성 물질인 Endothelin-1을 제거할 수 있는가?)

  • Chang, Yun-Hee;Ban, Ji-Eun;Lee, Hyoung-Doo;Lee, Sun-Hee;Rhym, Byuong-Yong;Sung, Si-Chan
    • Journal of Chest Surgery
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    • v.40 no.4 s.273
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    • pp.247-255
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    • 2007
  • Background: Lung injury that follows bypass has been well described. It is manifested as reduced oxygenation and lung compliance and, most importantly, increased pulmonary vascular resistance reactivity; this is a known cause of morbidity and mortality after repair of congenital heart disease. Injury to the pulmonary vascular endothelium, and its associated alterations of endothelin-1, is considered to be a major factor of bypass-induced lung injury. Removing endothelin-1 after bypass may attenuate this response. This study measured the concentration of serum and peritoneal effluent endothelin-1 after performing bypass to determine if endothelin-1 can be removed via peritoneal dialysis. Material and Method: From March 2005 to March 2006, 18 patients were enrolled in this study Peritoneal catheters were placed at the end of surgery. Serum samples were obtained before and after bypass, and peritoneal effluents were obtained after bypass. Endothelin-1 was measured by enzyme linked immunosorbent assay (ELISA). Result: In the patients with a severe increase of the pulmonary artery pressure or flow, the mean preoperative plasma endothelin-1 concentration was significantly higher than that in the patients who were without an increase of their pulmonary artery pressure or flow (4.2 vs 1.8 pg/mL, respectively, p<0.001). The mean concentration of plasma endothelin-1 increased from a preoperative value of $3.61{\pm}2.17\;to\;5.33{\pm}3.72 pg/ml$ immediately after bypass. After peritoneal dialysis, the mean plasma endothelin-1 concentration started to decrease. Its concentration at 18 hours after bypass was significantly lower than the value obtained immediately after bypass (p=0.036). Conclusion: Our data showed that the plasma endothelin-1 concentration became persistently decreased after starting peritoneal dialysis, and this suggests that peritoneal dialysis can remove the circulating plasma endothelin-1.

Low Frequency Noise Induces Stress Responses in the Rat (흰쥐에서 저주파소음에 의한 스트레스 반응)

  • Choi, Woong-Ki;Lee, Kyu-Sop;Joung, Hye-Young;Lee, Young-Chang;Sohn, Jin-Hun;Lee, Bae-Hwan;Pyun, Kwang-Ho;Shim, In-Sop
    • Science of Emotion and Sensibility
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    • v.10 no.3
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    • pp.411-418
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    • 2007
  • Exposure to low frequency noise(LFN) can lead to vibroacoustic diseases(VADs), which include a systemic disease with lesions in a broad spectrum of organs and a psychiatric condition. It is known that VAD is an established risk factor for the development of many psychological conditions in humans and rodents, including major depression and anxiety disorder. The present study investigated the effects of LFN on neuronal stress responses in the rat brain. The neuronal expression of the proto-oncogene c-fos in the paraventricular nucleus(PVN) of the hypothalamus and tyrosine hydroxylase(TH) in the LC was observed. The immunocytochemical detection of the Fos protein and TH has been used as a marker of neuronal activation in response to stress. In addition, corticosterone concentration was evaluated by using an enzyme-linked immunosorbent assay(ELISA). The LFN groups were exposed to 32.5Hz and 125Hz of noise(4hr/day for 2days). The numbers of c-fos and TH-immunoreactive cells in the PVN and LC were significantly increased in the LFN groups(32.5Hz and 125Hz) compared to the normal group. Corticosterone concentration in plasma was also increased in LFN groups. The present results demonstrated that exposure with LFN produced a pronounced increase in expression of c-Fos and TH in stress-relevant brain areas. These results suggest that the neural characteristics involved in LFN are similar to those activated by typical processive stressors. These results also suggest that the central and peripheral activations by LFN may be related to LFN-related negative behavioral dysfunctions such as VADs.

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Ehrlichia and Borrelia spp. Infection in German Shepherd Dogs in Korea (독일 셰퍼드 개에서 Ehrlichia와 Borrelia spp.의 감염)

  • Hwang, Cheol-Yong;Seo, Sang-Ho;Kang, Jun-Gu;Youn, Hwa-Young;Chae, Joon-Seok
    • Journal of Veterinary Clinics
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    • v.28 no.2
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    • pp.204-210
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    • 2011
  • The presence of the tick-borne pathogens Ehrlichia and Borrelia in German Shepherd dogs in Korea was determined by enzyme-linked immunosorbent assay (ELISA) and polymerase chain reaction (PCR). A total of 291 dogs were randomly selected from five Korean provinces from October 2005 through September 2006. The seroprevalence of antibodies to canine Ehrlichia and Borrelia agents detected by ELISA (Snap$^{(R)}$ 3Dx$^{(R)}$ Test, IDEXX Laboratories) was 7.56% (22 dogs) and 1.72% (5 dogs) respectively, throughout the country. Positive antibodies against both pathogens were detected in two dogs (0.69%). The provincial distribution of seroprevalence against Ehrlichia was 1.28% (1 of 78) in Gyeonggi-do, 12.64% (11 of 87) in Gangwon-do, 9.76% (4 of 41) in Chungcheong-do, 8.93% (5 of 56) in Gyeongsang-do, and 3.45% (1 of 29) in Jeolla-do. According to PCR analysis, Ehrlichia chaffeensis target DNA was amplified in 3.09% (9 of 291 dogs) of blood samples, 2.41% (7 of 291) from Gangwon-do and 0.69% (2 of 291) from Chungcheong-do. The oligonucleotide sequences (SNU-EC3 and SNU-EC5) from the PCR fragment examined in Korea were closely related to E. chaffeensis isolated from the tick Haemaphysalis longicornis, in China and the state of Arkansas in the US. Based on these results, the presence of E. chaffeensis infection was identified in German Shepherds being bred in Korea. These results bring to light the importance of paying close attention to tick-borne infections such as Lyme disease during clinical diagnosis. This infectious disease should be included as a differential diagnosis for patients who participate in outdoor activity from spring to fall or who have thrombocytopenia or leucopenia.

Regulatory Effect of Inflammatory Cytokines Secretion and Hypoxia-inducible $Factor-1{\alpha}$ Activation by Panax ginseng (인삼의 염증성 사이토카인 분비 및 저산소 유도인자-1${\alpha}$ 활성화 조절 효과)

  • Zo, Chul-Won;Lee, Seung-Hee;Kim, Dong-Woung;Lee, Seong-Kyun;Song, Bong-Keun
    • The Journal of Internal Korean Medicine
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    • v.27 no.4
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    • pp.864-878
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    • 2006
  • Purpose : Panax ginseng(PG) is considered to have salutary effects and stimulant actions on physical capacity. However, the effects of PG on the inflammatory cytokine secretion and hypoxia condition are still not understood. This study wasto elucidate the effect of PG on inflammatory cytokine secretion such as interleukin (IL)-1, IL-6, granulocyte macrophage colony stimulating factor (GM-CSF), and tumor necrosis factor $(TNF)-{\alpha}$. Also, the effects on the expression of vascular endothelial growth factor (VEGF) and hypoxia-inducible factor 1 (HIF-1) were measured. Methods : The water extract of PG was administrated to HMC-1 cells before phorbol myristate acetate (PMA)+A23187 treatment. $IL-1{\beta}$, IL-6, $TNF-{\alpha}$, GM-CSF, and VEGF secretion were measured by a modified enzyme-linked immunosorbent assay (ELISA). HIF-1 activation was measured by transcription factor enzyme-linked immunoassay (TF-EIA) Results : PG significantly decreased secretion of $IL-1{\beta}$, IL-6, $TNF-{\alpha}$, and GM-CSF in PMA+A23187-induced HMC-1 cells. VEGF secretion was not changed but HIF-1 activation was decreased by the treatment of PG. Conclusions : PG inhibited the secretion of inflammatory cytokines, which impliesPG might contribute to treatment of mast cell-mediated inflammatory disease. Also, PG inhibited PMA+A23187-induced $HIF -1{\alpha}activation}$ and DNA-binding activity for HIF-1. Therefore, these data demonstrate that PG modulates inflammatory cytokines through inhibition of $HIF-1{\alpha}activation}$ activation.

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Development of Assay Methods for Enterotoxin of Escherichia coli Employing the Hybridoma Technology (잡종세포종기법을 이용한 대장균의 장독소 측정법 개발)

  • Kim, Moon-Kyo;Cho, Myung-Je;Park, Kyung-Hee;Lee, Woo-Kon;Kim, Yoon-Won;Choi, Myung-Sik;Park, Joong-Soo;Cha, Chang-Yong;Chang, Woo-Hyun;Chung, Hong-Keun
    • The Journal of the Korean Society for Microbiology
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    • v.21 no.1
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    • pp.151-161
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    • 1986
  • In order to develop sensitive and sepcific assay methods for E. coli heat labile enterotoxin(LT) hybridoma cell lines secreting LT specific monoclonal antibody were obtained. LT was purified from cell lysate of E. coli O15H11. The steps included disruption of bacteria by French pressure, DEAE Sephacel ion exchange chromatography, Sephadex G200 gel filtration, and second DEAE Sephacel ion exchange chromatography, successively. Spleen cells from Balb/c mice immunized with the purified LT and $HGPRT^{(-)}$ plasmacytomas, $P3{\times}63Ag8.V653$ were mixed and fused by 50% (w/v) PEG. Hybrid cells were grown in 308 wells out of 360 wells, and 13 wells out of them secreted antibodies reacting to LT. Among these hybridoma cell 1G8-1D1 cell line was selected since it had produced high-titered monoclonal antibody continuously. By using culture supernatant and ascites from 1G8-1D1 cells the monoclonal antibody was characterized, and an assay system for detecting enterotoxigenic E. coli was established by double sandwich enzyme-linked immunosorbent assay (ELISA). The following results were obtained. 1. Antibody titers of culture supernatant and ascites from 1G8-1D1 hybridoma cells were 512, and 102, 400, respectively by GM1-ELISA and its immunoglobulin class was IgM. 2. The maximum absorption ratio of 1G8-1D1 cell culture supernatant to LT was 90% at $300\;{\mu}g/ml$ of LT concentration. LT concentration shown at 50% absorption ratio was $103.45{\mu}g$ and the absorption ratio was decreased with tile reduction of LT concentration. This result suggests that monoclonal antibody from 1G8-1D1 hybridoma cell bound with LT specifically. 3. The reactivities of 1G8-1D1 cell culture supernatant to LT and V. cholerae enterotoxin(CT) were 0.886 and 0.142(O.D. at 492nm) measured by the GM1-ELISA, indicating 1G8-1D1 monoclonal antibody reacted specifically with LT but not with CT. 4. The addition of 0.1ml of ascites to 0.6mg and 0.12mg of LT decreased the vascular permeability factor to 41% and 44% respectively, but it did not completely neutralize LT. 5. By double sandwich ELISA using monoclonal antibody, as little as 75ng of the purified LT per ml could be detected. 6. The results by assay of detecting LT in culture supernatants of 14 wild strains E. coli isolated from diarrhea patients by the double sandwich ELISA were almost the same level as those by reverse passive latex agglutination.

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Evaluation of the stability of IgM and specific antibody response of sevenband grouper Epinephelus septemfasciatus for application of antibody-detection ELISA (항체검출 ELISA 적용을 위한 능성어 IgM의 안정성 및 특이 항체 반응 평가)

  • Kim, Chun-Seob;Jang, Min-Seok;Kim, Wi-Sik;Kim, Jong-Oh;Kim, Du-Woon;Kim, Do-Hyung;Han, Hyun-Ja;Jeong, Sung-Ju;Oh, Myung-Joo
    • Journal of fish pathology
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    • v.22 no.3
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    • pp.335-342
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    • 2009
  • The stability of immunoglobulin M (IgM) on different serum storage conditions and specific antibody response were tested using the serum collected from sevenband grouper Epinephelus septemfasciatus by enzyme-linked immunosorbent assay (ELISA). To test the effect of storage temperature and duration, sevenband grouper antiserum against bovine serum albumin (BSA) was stored at -80, -20 or 4${^{\circ}C}$ for 1, 34, 61 or 119 days. In addition, to test the effect of repeated freeze-thawing condition, the anti-BSA fish serum was frozen at -20 and -80${^{\circ}C}$ and then thawn and frozen for 1, 5 or 10 times repeatedly. Consequently, no significant difference was found in ELISA optical density (O.D.) values of sera for the above mentioned storage conditions: different temperatures (-80, -20 and 4${^{\circ}C}$), durations of storage (1, 34, 61 and 119 days), and repeated thaw-freeze cycles (1, 5, and 10 times), indicating that IgMs of test fish were stable. The specific antibody response of sevenband grouper was observed after BSA-immunization of the test fish reared at 20 ${^{\circ}C}$ or 25${^{\circ}C}$. At the rearing temperature of 20${^{\circ}C}$, the specific antibody against BSA first appeared at 14 days and maximum antibody titer was observed between 21 and 28 days, while at the rearing temperature of 25 ${^{\circ}C}$, specific antibody appeared at 7 days and maximum antibody titer was observed between 14 and 21 days. In conclusion, the rearing temperature at 25${^{\circ}C}$ gave a faster and higher specific antibody response than at 20${^{\circ}C}$ and the specific antibody response maintained for approximately 2 months at 20℃ and 25${^{\circ}C}$.

Relationship between cord blood level of IL-12 in preterm newborns and development of wheezing (34주 미만 미숙아 제대혈청 내 interleukin-12 농도와 영아기 천명 발생과의 관계)

  • Yu, Hyo Jung;Park, Eun Ae;Kim, Ji Young;Cho, Soo Jin;Kim, Young Ju;Park, Hye Sook;Ha, Eun Hee
    • Clinical and Experimental Pediatrics
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    • v.51 no.7
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    • pp.754-759
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    • 2008
  • Purpose : Abnormal activation patterns of Th1/Th2-cells have been suggested to increase the prevalence of allergic diseases. Prevention is regarded as an important corner stone in the management of allergic diseases. In this study, we have investigated the relationship between cord blood levels of IL-4, IL-10, and IL-12 in preterm newborns and the development of allergic respiratory diseases in infancy Methods : Forty-six preterm newborns born at the Ewha Womans University Mokdong Hospital between January 2003 and July 2005, were enrolled for this study, and consent was obtained to test their cord blood samples. Clinical history was obtained from the hospital records. Cord blood was obtained at birth and kept frozen until it was tested. The levels of IL-4, IL-10, and IL-12 were determined by enzyme-linked immunosorbent assay (ELISA). Results : All infants were followed-up for a median of $16.0months{\pm}13.2d$ (range, 12.0 to 36.0 months). Eighteen infants who developed wheezing showed lower cord blood levels of IL-12 ($366.60{\pm}140.40$ vs $435.09{\pm}91.20pg/mL$, P=0.009). Cord blood levels of IL-4 and IL-10 showed no significant difference between the two groups. Four newborns who later developed asthma, and infants with asthma showed lower IL-12 level in the cord blood than other groups. Conclusion : Lower concentration of cord blood levels of IL-12 in newborns who later developed wheezing and asthma suggested that they had abnormal activation patterns of Th1/Th2-cells at the time of birth, and cord blood IL-12 level can be used as a predictor of allergic respiratory diseases.

The Seroprevalence and Related Factors of Helicobacter pylori Infection (Helicobacter pylori 감염의 유병률과 관련요인에 관한 연구)

  • Kim, Yeung-Wook;Lee, Su-Ill;Cho, Byung-Mann;Koh, Kwang-Wook;Kim, Young-Sil;Kang, Su-Yong;Cha, Oae-Ri;Kim, Don-Kyoun
    • Journal of Preventive Medicine and Public Health
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    • v.29 no.3 s.54
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    • pp.669-678
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    • 1996
  • Helicobacter pylori is now recognized as causative agent of chronic gastritis and peptic ulcer, and strongly associated with development of gastric carcinoma. With development of sensitive and specific serologic tests to identify individuals infected with Helicobacter pylori, the epidemiologic study of this diseases has been investigated. But it's transmission route is not established, yet. The purpose of this study is to measure the prevalence of Helicobacter pylori infection in healthy children and young adults and to evaluate related factors for Helicobacter pylori infection in Korea. The seroprevalence of Ig G antibodies to Helicobacter pylori was determined using a Enzyme Linked Immunosorbent Assay and we obtained the information, such as demographic characteristics, monthly household income, numbers of family members in the house, numbers of persons using same room, type of house, and type of drinking water through the questionnaire survey. The observed overall seropositivity rate was 25.7%. The rate is increased progressively from 5.8% in the age group $1\sim3$ years to 44.4% in the age group $20\sim29$years($\chi^2$ for trend, p<0.001). Especially, the rate increased steeply from 6.5% in the age group $4\sim6$ years to 20.8% in the age group $7\sim9$ years, and this suggested that elementary school age was the major acquisition time of Helicobacter pylori infection. In multivariate logistic regression model, age, numbers of family members in the house, and type of house was statistically significant variables for Helicobacter pylori infection. Each odds ratio(93% CI) were as follows; base to age group $1\sim9$ years, age group $10\sim19$ years $3.6(2.0\sim6.4)$, age group $20\sim29$ years $7.3(4.1\sim13.1)$ and base to group of $1\sim3$ family members, group of $4\sim5$ family members $2.1(1.1\sim4.0)$, group of 6 or more family members $2.7(1.3\sim5.4)$ and base to apartment, single and multihouse $1.9(1.1\sim3.5)$. Sex, monthly household income, numbers of persons using same room, and type of drinking water was not statistically significant for Helicobacter pylori infection.

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The Change of Serum Soluble E-selectin in Kawasaki Disease (가와사끼병에서의 혈청 Soluble E-selectin의 변화)

  • Jeong, Jae-Ho;Cho, Eun-Young;Lim, Jae-Woo;Cheon, Eun-Jeong;Ko, Kyong-Og;Lee, Kyung-Il
    • Clinical and Experimental Pediatrics
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    • v.48 no.5
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    • pp.539-544
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    • 2005
  • Purpose : The aim of this study was to investigate the pathophysiologic role of serum E-selectin, vascular endothelial growth factor(VEGF)-induced cell adhesion mollecule in Kawasaki disease(KD) and to look for the evidence of direct relationship between the plasma levels of soluble E-selectin and the incidence of coronary artery lesion(CAL). Methods : Changes in plasma levels of sE-selectin(n=98) over time were measured by enzyme-linked immunosorbent assay(ELISA) in 23 patients with acute KD and 25 age-matched febrile children. Results : Compared with control values, the peak levels of plasma sE-selectin were significantly elevated($mean{\pm}S.E$. : $22.89{\pm}12.53ng/mL$ vs $10.65{\pm}3.42ng/mL$, P=0.01) in KD. 5 patients with CAL, plasma sE-selectin levels before treatment were higher than in 18 patients without CAL($mean{\pm}S.E$. : $39.43{\pm}15.08ng/mL$ and $19.00{\pm}8.32ng/mL$, respectively; P=0.01). Plasma sE-selectin declined rapidly in the majority of KD patients regardless of the presence of CAL. Plasma sE-selectin levels after treatment and convalesent period were similar in KD patients with and without CAL. The plasma levels sE-selectin were correlated with those of white blood cell count(r=0.299, P<0.05), CRP(r=0.430, P<0.05), serum albumin(r=-0.483, P<0.05), serum protein(r=-0.502, P<0.05) and hemoglobin(r=-0.372, P<0.05) not with those of ESR, platelet, or duration of fever. There were significant differences in the initial level of serum sE-selectin between KD with and without CAL($mean{\pm}S.E$. : $39.44{\pm}15.08ng/mL$ vs. $19.00{\pm}17.18ng/mL$) in multivariated linear tests. Conclusion : Plasma sE-selectin levels were significantly higher in KD than in other febrile illness. Higher plamsa levels of sE-selectin may have potential as a predictor of CAL in patients with KD.