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Sperm-Mediated Gene Transfer by Injection of Sperm or Sperm Head into Porcine Oocytes

  • S.Y. Ahn;Lee, H.T.;K. S. Chung
    • Proceedings of the KSAR Conference
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    • 2001.03a
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    • pp.56-56
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    • 2001
  • The exogenous gene transfer by intracytoplasmic sperm injection (ICSI) procedure has been recently used to produce transgenic mice and pigs. Sperm-mediated DNA transfer has the potential to markedly simplify the generation of transgenic animals. This method may serve as an alternative to the pronucleus injection of DNA for the production of transgenic pigs. Therefore, in this study, we investigated the expression of transgene after co-injection of spermatozoon or sperm head with green fluorescent protein (GFP) gene into in vitro matured porcine oocytes. Spermatozoon and sperm head, that was obtained by sonication, were treated with 0.03% Triton X-100 to remove the membrane. They were preincubated with linearized pEGFP-N1 for 1 min, and then embryos cultured NCSU23 medium for 2.5 days after co-injected of sperm and DNA. We monitored expression of GFP in embryos under epifluorescent microscope. The remove of sperm membrane did not alter the developmental competence of embryos after ICSI. At 7 days following injection, the rates of blastocysts following injection of intact sperm (15.0%), and of sperm with disrupted membrane (14.2%) were higher than that following IVF (10.0%). Porcine oocytes injected with sperm which co-cultured with DNA concentration of 1, 0.1, and 0.01 ng were 60, 65.7 and 75% and 18.5, 37.4 and 22.2% for rates of cleavage and GFP expression, respectively. In vitro matured porcine oocytes injected with sperm and isolated sperm head resulted in 69 and 59.7% of cleavage rates, respectively The rates of embryo GFP expressed did not significantly different between sperm (20.4%) and sperm head (20.0%) injection. The transgenic embryos with the clusters of positive blastomeres were observed under fluorescent microscope. Most of embryos expressed GFP gene showed mosaicism. They showed GFP expression at 1/4, 2/4 and 3/4 of blastomeres at the 4-cell stage. Among these 4-cell embryos, the expression rate of 1/4 blastomere group (54.6%) was higher than the other groups (15.3-30.7%). These results indicate that membrane disrupted sperm could attach with exogenous DNA, and that this procedure may be useful to introduce foreign gene into porcine oocytes. Therefore, our data suggest that the ICSI car be a useful tool to efficiently produce transgenic pig as well as other mammals.

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A Study on Nutritional Status of the Long-lived Elderly People in Kyungnam (경남 일부지역 장수노인의 영양섭취상태에 관한 연구)

  • 최희정;김경업;김성희;강동희;정효숙
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.31 no.5
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    • pp.877-884
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    • 2002
  • This study was carried out to examine nutrients intake and food preference in 100 elderly people aged over 85 years residing in Namhae-gun Kyungnam. Mean daily energy intake and Korean recommended dietary allowances (RDA) percent were 1547.6$\pm$452.7 kcal and 91.5%, respectively. Mean daily intakes of protein (63.1$\pm$28.2 g), iron (16.7$\pm$11.2 mg), vitamin Bi (1.0410.4 mg) and vitamin C (97.0$\pm$45.3 mg) were higher than RDA. The intakes of calcium (418.1$\pm$189.7 mg) and vitamin B2 (0.94$\pm$0.5 mg) were shown to be 58.4% and 78.4% of the RDA. Females had higher intakes of energy and most other nutrients than males. Major food source of energy and protein intakes was rice and cereals, and that of lipid intake was meat, fish, egg and beans. Cooked rice, all kind of vegetables, fish, yogurt and sugars were preferred to the subjects. The most preferred taste was sweet.

Removal of Chromium (VI) by Escherichia coli Cells Expressing Cytoplasmic or Surface-Displayed ChrB: a Comparative Study

  • Zhou, Xiaofeng;Li, Jianghui;Wang, Weilong;Yang, Fan;Fan, Bingqian;Zhang, Chenlu;Ren, Xiaojun;Liang, Feng;Cheng, Rong;Jiang, Fengying;Zhou, Huaibin;Yang, Juanjuan;Tan, Guoqiang;Lyu, Jianxin;Wang, Wu
    • Journal of Microbiology and Biotechnology
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    • v.30 no.7
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    • pp.996-1004
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    • 2020
  • Various genetically engineered microorganisms have been developed for the removal of heavy metal contaminants. Metal biosorption by whole-cell biosorbents can be enhanced by overproduction of metal-binding proteins/peptides in the cytoplasm or on the cell surface. However, few studies have compared the biosorption capacity of whole cells expressing intracellular or surface-displayed metal-adsorbing proteins. In this study, several constructs were prepared for expressing intracellular and surface-displayed Ochrobactrum tritici 5bvl1 ChrB in Escherichia coli BL21(DE3) cells. E. coli cells expressing surface-displayed ChrB removed more Cr(VI) from aqueous solutions than cells with cytoplasmic ChrB under the same conditions. However, intracellular ChrB was less susceptible to variation in extracellular conditions (pH and ionic strength), and more effectively removed Cr(VI) from industrial wastewater than the surface-displayed ChrB at low pH (<3). An adsorption-desorption experiment demonstrated that compared with intracellular accumulation, cell-surface adsorption is reversible, which allows easy desorption of the adsorbed metal ions and regeneration of the bioadsorbent. In addition, an intrinsic ChrB protein fluorescence assay suggested that pH and salinity may influence the Cr(VI) adsorption capacity of ChrB-expressing E. coli cells by modulating the ChrB protein conformation. Although the characteristics of ChrB may not be universal for all metal-binding proteins, our study provides new insights into different engineering strategies for whole-cell biosorbents for removing heavy metals from industrial effluents.

A clinical study on the etiology of parapneumonic effusion in children (소아 감염성 흉막삼출의 원인 분석)

  • Yeom, Jung-Sook;Bae, Won-Tae;Park, Eun-Sil;Seo, Ji-Hyun;Lim, Jae-Young;Park, Chan-Hoo;Woo, Hyang-Ok;Youn, Hee-Shang
    • Clinical and Experimental Pediatrics
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    • v.49 no.1
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    • pp.56-63
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    • 2006
  • Purpose : This study was designed to document the etiologies and the characteristics of parapneumonic effusion in children. Methods : During a 17-year period from 1987 to 2004, parapneumonic effusion was confirmed in 86 children at Gyeongsang National University Hospital. The clinical records of these children were reviewed and radiological findings and laboratory data, especially results of thoracentesis, were analyzed retrospectively. Results : M. pneumoniae(34 subjects) was the most common pathogen at all over age, especially above 1-years-old. There were diagnosed with clinical characteristics and serologic tests. The $2^{nd}$ most common pathogen revealed non tuberculous bacteria(14 subjects). A species of bacteria at no tuberculous bacteria revealed S. aureus(5), S. pneumoniae(3), P. aeroginosa(3), other staphylococcus (2), and K. pneumoniae(1). There were confirmed with sputum culture or pleural fluid culture or blood culture. S. aureus was most common pathogen in infants. The $3^{rd}$ common pathogen was M. tuberculosis(7). There were confirmed with skin tuberculin tests and AFB stains. Another that was classified as a non bacteria was adenovirus(2). Complications of parapneumonic effusion such as pleural thickness occurred on M. tuberculosis(1). Non tuberculous bacteria, especially S. aureus revealed a serious predominance of polymorphocyte at pleural fluid, and lowest pleural pH and glucose, and highest pleural protein and LDH. Tuberculosis revealed high pleural protein and LDH. Conclusion : Age and chemistries of pleural fluid might be helpful in differentiating various etiologies of parapneumonic effusion. If there were suspicious of tuberculosis and non-tuberculous bacteria, more aggressive approaches were needed to prevent complication.

In Vitro Phosphorylation of Nuclear Proteins in Isolated Liver Nuclei from Rats Maintained in a Starvation State, Following Refeeding, and from Diabetic Rats with Insulin Injection (단식(斷食), 재급식(再給食) 및 인슈린 투여(投與) 후(後)에 쥐의 간(肝)으로부터 분리된 세포핵의 핵단백질 인산화)

  • Lee, Hyo-Sa;Gibson, David M.
    • Applied Biological Chemistry
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    • v.23 no.1
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    • pp.23-30
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    • 1980
  • Labelling of chromatin proteins with 32P was observed after incubating isolated liver nuclei with $[{\gamma}-32P]$ ATP for 5 minutes at $37^{\circ}C$. The pattern of labelling with 32P was examined on SDS polyacrylamide gel electrophoresis with nuclei from rats maintained in a starvation state for 48 hours, following refeeding for 12 hours; and from fed streptozotocin-diabetic rats with insulin injection 6 hours before sacrifice. With 48h starved rat liver nuclei the level of phosphorylation for 0.14M NaCl soluble proteins was decreased in the molecular weights between 41,000 and 200,000 daltons relative to normal controls. Refeeding the starved rats reversed the change of phosphorylation pattern over 12 hour The level of phosphorylation for five phenol soluble non-histone proteins with molecular weights above 59,000 daltons was somewhat decreased with 48h starved rat liver nuclei as compared with that of normal controls. Starvation also decreased the phosphorylation level of major histones in relation to normal controls. The experiment with insulin injection into fed streptozotocin-diabetic rats showed the tendency to increase phosphorylation of 0.14M NaCl soluble proteins (130,000 dalton protein) and phenol soluble non-histone proteins (155,000 dalton protein). The phosphorylation level of histones appeared to be invariant under the experimental conditoins employed here. These results suggest the possibility that the phosphorylation and dephosphorylation of 0.14M NaCl soluble proteins and $H_1$ histone precede those of other chromatin associated nuclear proteins, It is of interest to find that insulin signal was correlated to phosphorylation of nuclear proteins while glucagon signet dephosphorylated nuclear proteins.

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Study on Characteristic of Pyropia Dentata Three Cultivar in Haenam Aquafarm, Jellanam-do (전남 해남에서 잇바디돌김 3종의 양식 특성 연구)

  • Han, M.K.;Jeong, D.S.;Kim, C.W.;Choi, S.J.
    • Journal of Practical Agriculture & Fisheries Research
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    • v.23 no.2
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    • pp.43-50
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    • 2021
  • Growth tests on the Yuldo, Supum1 and Supum2 cultivars of Pyropia dentata were performed at the Eoran and Imha aquafarm, Haenam in Jeollanamdo, from October to December in 2017. The mean water temperature ranged from 5.4 to 26.4 ℃. In Eoran aquafarm (flating raft method), dissolved total nitrogen (DTN), dissolved total phosphorus (DTP), total nitrogen (TN), total phosphorus (TP) and chemical oxygen demand (COD) ranged from 0.091 to 0.181 mg/L, 0.007 to 0.019 mg/L, 0.114 to 0.187 mg/L, 0.008 to 0.033 mg/L and 0.200 to1.000 mg/L, respectively. In Imha aquafarm (fixed pold method), DTN, DTP, TN, TP and COD ranged from 0.118 to 0.276 mg/L, 0.005 to 0.024 mg/L, 0.155 to 0.305 mg/L, 0.009 to 0.042 mg/L and 0.300 to1.400 mg/L, respectively. In order to investigate the number of conchospores attached on the Pyropia net, which was cut into about 4cm long. The mean number of conchospores of Yuldo, Supum1 and Supum2 cultivars were 39.5, 26.5, 72.5, respectively. Young thalli were harvested two times at Eoran aquafarm, and three times at Imha aquafarm. In eoran aquafarm, the mean thallus length of Yuldo, Supum1 and Supum2 cultivars were 49.9, 46.0, 42.0 cm on October and 163, 126.0, 263.0 cm on November, respectively. The mean thallus width of Yuldo, Supum1 and Supum2 cultivars were 5.8, 4.6, 11.5 cm on October and 20.9, 11.5, 14.0 cm on November, respectively. In Imha aquafarm, the mean thallus length of Yuldo, Supum1 and Supum2 cultivars were 119.0, 60.9, 71.0 cm on October, 196.0, 132.0, 262.0 cm on November and 187.0, 281.0, 296.0 cm on December, respectively. The mean thallus width of Yuldo, Supum1 and Supum2 cultivars were 4.2, 3.4, 3.1 cm on October, 8.9, 6.2, 6.6 cm on November and 11.7, 11.6, 9.4 cm on December, respectively. In eoran aquafarm, contents of moisture, crude ash, crude lipid, crude protein and carbohydrate of three cultivars ranged from 11.64 to 20.15, 19.54 to 21.19, 0.00 to 0.18, 29.78 to 37.81, 29.16 to 29.71, respectively. In Imha aquafarm, contents of moisture, crude ash, crude lipid, crude protein and carbohydrate of three cultivars ranged from 8.43 to 9.15, 11.42 to 17.49, 0.00 to 0.00, 31.90 to 37.54, 36.30 to 42.24, respectively.

A Survey of Nutritional Status on Pre-School Children in Korea (학영기전아동(學齡期前兒童)의 영양실태조사(營養實態調査))

  • Ju, Jin-Soon;Oh, Seoung-Ho
    • Journal of Nutrition and Health
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    • v.9 no.2
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    • pp.68-86
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    • 1976
  • The primary purpose of this study is to evaluate the correct nutritional status on pre-school children in Korea. Furthermore, it made an attempt to find and define nutrional problems, and assist in establishment on their nutritional improvement plan. For this, food intake and health condition (physical, clinical, biochemical and parasitological) survey on 109 Pre-school children in both sexes, randomly selected from Yang-Gu area in Gang-Won province and Rea-ju area in Kyong-gy Province, were conducted by means of three-day records, during the two periods of Spring and Fall season in 1975. The results obtained are summerized as follows: 1. The food intake; Average food intake of the subjects per day were $508{\sim}647g$ ($83{\sim}91%$ in vegetable foods and $5.5{\sim}11.7%$ in animal foods) in Yang-gu area, and $587{\sim}698g$ ($88{\sim}89%$ in vegetable foods and $6.3{\sim}7.6%$ in animal foods) in Rea-ju area. 2. The intake of energy and nutrients; a) Calory intake. Average energy intake of subjects per day in Yang-gu area$(1120{\sim}1415kcal)$ were all lower than the Korean Recommended Dietary Allowances (RDA) in either Spring and Fall survey, whereas the subjects in Rea-ju area were lower intake $(1213{\sim}1418kcal)$ than the RDA in the Spring but higher intake$(1516{\sim}1755kcal)$ than the RDA in the Fall, and the average intake were similar level with that of RDA. b) Protein intake. Average protein intake of the subjects per day in Yang-gu area $(33{\sim}43g)$ girl subjects in Rea-ju area $(35{\sim}39g)$ were lower than the RDA in either Spring and Fall survey, whereas the boy subjects in Rea-ju area$(36{\sim}38g)$ were lower in Spring and higher $(49{\sim}57g)$ in the Fall than that of the RDA, but the average $(43{\sim}47g)$ were similar level with the RDA. The protein intake from animal sources in all subjects were much lower $(5.5{\sim}11.7\;of\;total\;protein)$ than the RDA. c) Fat intake. Average fat intake were very lower in all subjects of both area $(14{\sim}24g\;in\;Yang-gu,\;10{\sim}12g\;in\;Rea-ju)$ than that of RDA which is recommended $12{\sim}14%$ of total energy to be supplied from fat. d) Calcium intake. Average calcium intake were very low in all subjects of both area $(264{\sim}355mg\;in\;Yang-gu\;and\;283{\sim}429mg\;in\;Rea-ju)$, especially, these in Spring were about a half level of the RDA, and it was much increased in the Fall due to increased intake of milk, but it was still not enough than the RDA. e) Vitamin A intake. Average intake of V.A ($703{\sim}1465\;IU$ in Ynag-gu and $750{\sim}1521\;IU$ in Rea-ju) were also lower than the RDA, moreover their V-A sources were mainly vegetable, so that the V-A supply might be critical one for the subjected. f) Riboflavin intake. Average riboflavin intake on all subjects in both area except boys in Rea-ju area in Fall, were very lower than the RDA. 3. The physical status; a) Average weight and height of boys aged 4 and 5 in Yang-gu area and girls of aged 5 in Rea-ju area were lower than those of Korean Standard of 1967 report, but those by age of girls in Yang-gu area and boys in Rea-ju area were a little heigher than the Korean Standard. It is, hower, present Korean standard of physical status might be somehow heigher than the 1967, since the socio-economical situation has been much improved during past a decade. So that, if one considered on this sense, the physical status of the subjects on this survey might be somehow lower than those of present Korean standard. b) Average upper arm circumference in both area were no difference each other, and their mean values of age 4, 5 and 6 in boy and girl were 15.6, 16.5, 16.4 and 15.5, 16.5, 16.4cm respectively. c) Average chest girth of boys were similar to those of Korean standard whereas the girls were smaller than the Korea standard. The average head circumference also showed similar tendency with the chest girth. 4. The clinical findings; The most popular clinical signs were angular stomatitis and dental caries, and boys had more heigher incidence then the girls. 5. The biochemical findings; a) Hemoglobin and anemia Average Hb value of boys and girls were 11.4 and 10.9g per 100 ml of blood respectively. The incidence of anemia (Hb value below 11 g/100 ml, by WHO) was increased by age, and girls had more heigher incidence than the boy (34% : 48%). The incidence of anemia in age of 4,5, and 6 in boys and girls were 28%, 41% 34%, and 33%, 50%, 49% respectively. The degree of the anemia was not severe, and the anemia of there subjects may be caused mainly low intake of better quality protein and low iron intake as well. b) Hematocrit. Average Ht value of whole subject were $39.9{\sim}41.6%$. c) Blood plasma protein. Average blood plasma protein contents of whole subjects were $6.6{\sim}7.4gm$ per 100 ml. The incidence of deficient range (<6.0g%, by ICNND) was only one girl of age 4 in yang Gu area. 6. Parasitological findgs; The most popular parasitism were asicris lumbicoides and trichocephalus trichiura, and about 2/3 of the whole subjects were suffering one or more of these parasitism.

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Comparisons of Characteristics of Amaranth Starches Isolated from five Cultivars Grown in Korea (국내산 아마란스로부터 분리한 전분의 특성 비교)

  • Choi, Cha-Ran;Choi, Hun-Jae;Kim, Sung-Ran;Lee, Jae-Hak;Shin, Mal-Shick
    • Korean Journal of Food Science and Technology
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    • v.32 no.2
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    • pp.252-257
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    • 2000
  • The physicochemical and pasting properties of amaranth starches isolated from five cultivars, Andy, Suvarna, Nu World, K266-1, K432, grown in Korea, were investigated. The shapes of starch granules were all polygonal and size was in the range $1.14{\sim}1.48\;{\mu}m$ for all five cultivars. X-ray diffractograms were shown typical A type diffraction patterns for all amaranth starches. The protein and crude lipid contents of starches were $0.13{\sim}0.23%$ and $0.01{\sim}0.05%$, respectively. The apparent amylose contents ranged from $2.79{\sim}4.35%$ and the water binding capacities were $128.05{\sim}135.80%$. The transmittances of 0.1% amaranth starch suspensions except K266-1 increased rapidly above $65^{\circ}C$, thereafter increased slowely. The initial pasting and peak temperature ranges of five cultivars by RVA were $71.3{\sim}73.7^{\circ}C$ and $81.5{\sim}84.0^{\circ}C$, respectively. The peak and cooling viscosities followed the order : Nu World>Andy>Suvarna>K432>>K266-1. Nu World was shown the highest peak (166), final (103) and cooling viscosities (30 RVU). K266-I exhibited the lowest setback (-38) of all five cultivars. Gelatinization (To) and peak temperature (Tp) of amaranth starches in DSC thermograms were $65.7{\sim}68.0^{\circ}C$ and $70.6{\sim}75.8^{\circ}C$, respectively. Enthalpies followed the order: K266-1

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Growth Characteristics and Forage Productivity of New Forage Barley Variety, 'Miho' (청보리 신품종 '미호'의 생육특성과 수량성)

  • Oh, Young-Jin;Park, Tae-Il;Park, Hyoung-Ho;Han, Ouk-Kyu;Song, Tae-Hwa;Park, Jong-Chul;Kim, Yang-Kil;Park, Jong-Ho;Kang, Hyeon-Jung;Kang, Chon-Sik;Cheong, Young-Keun;Kim, Kyong-Ho;Kim, Bo-Kyeong;Yun, Geon-Sig;Hong, Gi-Heung;Bae, Jeong-Suk;Lee, Seong-Tae
    • Journal of The Korean Society of Grassland and Forage Science
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    • v.36 no.4
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    • pp.370-375
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    • 2016
  • The purpose of development new variety 'Miho' (Hordeum vulgare L.) is a favorite with livestock feed and develop varieties resistant to disease and lodging. 'Miho' was carrying out the growth habit of group III with green and mid-wide leaf. Awn that is related to preference of livestock is a semi-smooth awn. This cultivar had 96 cm of culm length, 650 of spikes $per\;m^2$. Heading date of 'Miho' was April 27, and maturing date on May 30, which were later than cultivar 'Youngyang'. It also showed strong winter hardiness, and similar resistance to shattering and bariy yellow mosaic virus (BaYMV) compared with those of check one. The best thing among the traits of a good quality with the plant green at the latter growing period. The average forage dry matter yield in the regional yield trial was about 13.1, 12.1 MT per ha in upland and paddy field, respectively, which were 9%, 2% higher than that of the check cultivar. It's also showed 6.8% crude protein, 27.1% ADF (acid detergent fiber), and 67.5% TDN (total digestible nutrients), including higher silage quality for whole crop barley. This cultivar would be suitable for the area whose average minimum temperature was above $-8^{\circ}C$ January in Korean peninsula.

Analysis of antigen specificity using monoclonal and polyclonal antibodies to cysticercus cellulosae by enzyme-linked immunoelectrotransfer blot technique (효소면역전기영동이적법을 이용한 유조설고충 단세후군항체 및 환기혈청에 대한 항원특리성 분석)

  • Jo, Seung-Yeol;Gang, Sin-Yeong;Kim, Seok-Il
    • Parasites, Hosts and Diseases
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    • v.25 no.2
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    • pp.159-167
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    • 1987
  • To analyse the antigen specificity of patients sera from 24 confirmed neurocysticercosis and a monoclonal antibody, SDS-PAGE using 10~15% linear gradient gel and EITB were done. Cystic fluid, saline extracts of scolex and of whole worm of C. cellulosae, saline extracts of sparganum, hydatid cyst fluid, saline extracts of Fasciola, Clonorchis and Paragonimus were used as antigen. Of protein bands in cystic fluid of C. cellulosae, patient sera reacted frequently to bands of 152, 94, 64, 48, 24, 15, 10 and 7kDa proteins. To saline extracts of scolex and whole worm of C. cellulosae, patients sera reacted frequently to 94, 64, 52, 39, 34, 15 and 10kDa bands. Two bands in sparganum extract (130 and 64kDa) and two bands in hydatid cyst fluid (52 and 27kDa) were cross-reacting bands with sera from cysticercosis patients. Saline extracts of Fasciola, ClonorchiJ and Paragonimus did 'not exhibit cross-reacting bands. Monoclonal antibody to cystic fluid of C. cellulosae was found to react with low molecular weight proteins of 15, 10 and 7kDa.

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