• Title, Summary, Keyword: FITC conjugate

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Specific Targeting of Fluorescein Isothiocyanate with Ep-CAM Antibody(Specific targeting of FITC with Ep-CAM Antibody)

  • Lee, Young-Tae;Tae, Gun-Sik
    • Journal of Photoscience
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    • v.10 no.3
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    • pp.237-240
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    • 2003
  • The tetradecameric peptide (K47-K60) near the NH$_2$-terminal region of epithelial-cell adhesion molecule (Ep-CAM) was chosen as antigenic site and a polyclonal antibody was generated, which could recognize Ep-CAM from the mouse colon tissue or the colon cancer cell, CT-26, in Western blot analysis. Then, the fluorescein isothiocyanate (FITC), a fluorescence dye, was conjugated with the affinity purified Ep-CAM antibody using thiocyanate and the amino groups of FITC and antibody, respectively. The molar ratio of FITC to antibody was estimated approximately 1.86 to 1.00 by measuring the optical densities at 492 nm and 280 nm. Ep-CAM antibody-FITC conjugate was then used for immunohistochemistry of the CT-26 cells. Judging from the shapes formed by fluorescence, the Ep-CAM antibody could delivered FITC to the surface of cells in which Ep-CAM was expressed. This result implies that Ep-CAM antibody could be also used for the tissue-specific delivery of the photosensitizer to the target protein via antigen-antibody interaction.

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Modulation of Electroosmotic Flow through Skin: Effect of Poly(Amidoamine) Dendrimers

  • Kim, Hye Ji;Oh, Seaung Youl
    • Biomolecules & Therapeutics
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    • v.26 no.2
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    • pp.182-190
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    • 2018
  • The objective of this work is to evaluate the effect of polyamidoamine (PAMAM) dendrimers on electroosmotic flow (EOF) through skin. The effect of size and concentration of dendrimer was studied, using generation 1, 4 and 7 dendrimer (G1, G4 and G7, respectively). As a marker molecule for the direction and magnitude of EOF, a neutral molecule, acetoaminophen (AAP) was used. The visualization of dendrimer permeation into the current conducting pore (CCP) of skin was made using G4-fluorescein isothiocyanate (FITC) conjugate and confocal microscopy. Without dendrimer, anodal flux of AAP was much higher than cathodal or passive flux. When G1 dendrimer was added, anodal flux decreased, presumably due to the decrease in EOF by the association of G1 dendrimer with net negative charge in CCP. As the generation increased, larger decrease in anodal flux was observed, and the direction of EOF was reversed. Small amount of methanol used for the preparation of dendrimer solution also contributed to the decrease in anodal flux of AAP. Cross-sectional view perpendicular to the skin surface by confocal laser scanning microscope (CLSM) study showed that G4 dendrimer-FITC conjugate (G4-FITC) can penetrate into the viable epidermis and dermis under anodal current. The permeation route seemed to be localized on hair follicle region. These results suggest that PAMAM dendrimers can permeate into CCP and change the magnitude and direction of EOF. Overall, we obtained a better understanding on the mechanistic insights into the electroosmosis phenomena and its role on flux during iontophoresis.

Viualization of Progesterone Binding to Plasma Membrane of Xenopus Oocytes

  • Ju, Jung-Won;Im, Wook-Bin;Kwon, Hyuk-Bang;Choi, Hueng-Sik
    • Animal cells and systems
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    • v.5 no.1
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    • pp.51-57
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    • 2001
  • We have previously shown that oocyte maturation is induced by an immobilized progesterone, progesterone-3-carboxymethyloxime - bovine serum albumin conjugate (P-BSA) in Rana dybowskii. In this study, we confirmed the maturation inducing activity of P-BSA on Xenopus oocyte and examined the binding character of the immobilized progesterone on the surface of Xenopus oocytes after removal of the vitelline layer. P-BSA induced maturation of Xenopus oocytes but E-BSA failed to do so as observed in Rana. Binding of the immobilized progesterone, fluorescein isothiocyanate-labeled progesterone-3-0-carboxymethyloxime-BSA (P-BSA-FITC) on the devitellined oocytes surface was examined by fluorescence confocal microscopy. The binding affinity of P-BSA-FITC to the devitellined oocyte was higher than that of estrogen-BSA-FITC (E-BSA-FITC) or testosterone-BSA-FITC (T-BSA-FITC). The binding disappeared in the presence of excess free progesterone but not in the presence of free estrogen. Maximum binding occurred after two-hours of incubation with P-BSA-FITC at pH 7.5. Stronger binding occurred in oocytes at stage Vl than stage IV, and in vitro treatment of hCG enhanced the binding. Taken together, these results suggest that a specific receptor for progesterone exists on the plasma membrane of Xenopus oocytes and that progesterone acts initially on this putative receptors and triggers generation of membrane-mediated second messengers during the early stage of oocyte maturation In amphibians.

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A Blood-brain Barrier Permeable Derivative of 5-Fluorouracil: Preparation, Intracellular Localization, and Mouse Tissue Distribution

  • Im, Jung-Kyun;Biswas, Goutam;Kim, Wan-Il;Kim, Kyong-Tai;Chung, Sung-Kee
    • Bulletin of the Korean Chemical Society
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    • v.32 no.3
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    • pp.873-879
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    • 2011
  • 5-Fluorouracil (5-FU), an anticancer agent was covalently attached to the recently developed sorbitol-based G8 transporter, and the conjugate (7) with FITC was found to have an affinity toward mitochondria and to readily cross BBB to gain an entry into mouse brain. Measured by $IC_{50}$, the conjugate (9) without the fluorophore showed enhanced cytotoxic activity toward two types of multidrug-resistant cell lines. These results strongly suggest that the sorbitol-based G8 transporter can be utilized as a good CNS delivery vector.

Transdermal Delivery of Bufexamac using Microneedle System (마이크로니들시스템을 이용한 부펙사막의 경피전달)

  • Mo, Jong Hyun;Kim, Yun Tae;Park, Jung Soo;Lee, Jun Hee;Lee, Tae-Wan;Khang, Gilson;Rhee, John Moon;Lee, Han-Koo;Lee, Hai Bang
    • Korean Chemical Engineering Research
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    • v.46 no.2
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    • pp.211-216
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    • 2008
  • Bufexamac which was used for treatment of atopic dermatitis is the drug which was made as the ointment. However, penetration rate of bufexamac was very low for the barrier effect of stratum corneum. Microneedle was used to increase transdermal delivery rate of the bufexamac. We tried to conjugate bufexamac and FITC for the detection of penetration rate of bufexamac. FITC-bufexamac was mixed in hydrogel for the treatment skin surface. Fluorescent spectrophotometer was used to analysis the concentration of FITC-bufexamac. Microscope using fluorescent filter was used to capture the image about location of FITC-bufexamac in the skin. We confirmed that permeation rate of bufexamac was increased with the treatment by microneedle and was increased by the increasing number treatment of microneedle.

Surface Modification of Glass Chip for Peptide Microarray (펩타이드 Microarray를 위한 유리 칩의 표면 개질)

  • Cho, Hyung-Min;Lim, Chang-Hwan;Neff, Silke;Jungbauer, Alois;Lee, Eun-Kyu
    • KSBB Journal
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    • v.22 no.4
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    • pp.260-264
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    • 2007
  • Peptides are frequently studied as candidates for new drug development. Recently, synthesized peptide library is screened for a certain functionality on a microarray biochip format. In this study, in order to replace the conventional cellulose membrane with glass for a microarray chip substrate for peptide library screening, we modified the glass surface from amines to thiols and covalently immobilized the peptides. Using trypsin-FITC (fluorescein isothiocyanate) conjugate that could specifically bind to a trypsin binding domain consisting of a 7-amino acid peptide, we checked the degree of surface modification. Because of the relatively lower hydrophilicity and reduced surface roughness, the conjugation reaction to the glass required a longer reaction time and a higher temperature. It took approximately 12 hr for the reaction to be completed. From the fluorescence signal intensity, we could differentiate between the target and the control peptides. This difference was confirmed by a separate experiment using QCM. Furthermore, a smaller volume and higher concentration of a spot showed a higher fluorescence intensity. These data would provide the basic conditions for the development of microarray peptide biochips.

Isolation of infectious pancreatic necrosis virus from rainbow trout in Korea (국내(國內) 무지개 송어(松魚)에서의 IPN 바이러스의 분리(分離))

  • Lyoo, Young-soo;Chang, Chung-ho;Jean, Young-hwa;Lee, Jong-oh;Rhee, Je-chin
    • Korean Journal of Veterinary Research
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    • v.31 no.2
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    • pp.195-199
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    • 1991
  • Infectious pancreatic necrosis(IPN) virus was known as a causative agent of newly recognized viral disease of young rainbow trout characterized by highly contagious, high mortality and necrosis of pancreas. Several strains of IPN viruses were recovered from young rainbow trout that have been shown a typical cinical sign of infectious pancreatic necrosis disease. The field isolate produced cytopathic effect, and multiplied up to $10^{6.0}$ to $10^{6.5}$ $TCID_{50}/0.1ml$ in BT cell culture. In the indirect immunofluorescent assay with trout anti-IPN virus IgG and goat anti-trout IgG FITC conjugate, these isolates were proved to be a IPN virus that were closely related with VR277 strain of IPN virus antigenically.

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Effect of Enterococcus faecalis strain PL9003 on Adherence and Growth of Helicobacter pylori

  • Nam, Hye-Ran;Ha, Mi-Sun;Lee, En-Jung;Lee, Yeon-Hee
    • Journal of Microbiology and Biotechnology
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    • v.12 no.5
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    • pp.746-752
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    • 2002
  • The purpose of the present study was to examine the antagonistic activities of Enterococcus faecalis strain PL9003 (PL9003) on Helicobacter pylori. This strain was isolated from infant feces and found to inhibit both the growth of H. pylori and its in vitro adherence to the human gastric cell line MKN-45. The binding of PL9003 to MKN-45 was observed under a light microscope after Cram staining and under a scanning electron microscope. When detected with an FITC-conjugate antibody, both viable and nonviable PL9003 were found to decrease the number of H. pylori bound to MKN-45. When detected by an enzyme-linked immunoabsorbent assay, about 70% of the H. pylori bound on MKN-45 disappeared with the four-1314 addition of viable or nonviable PL9003. The spent culture supernatant (SCS) of PL9003 also decreased the viability of H pylori even after neutralization and pepsin treatment. The above results suggest that PL9003 has a potential as a new probiotic for the stomach.

Surface Modification of Magnetites Using Maltotrionic Acid and Folic Acid for Molecular Imaging

  • Selim, K.M.Kamruzzaman;Lee, Joo-Hee;Kim, Sun-Jung;Xing, Zhicai;Kang, Inn-Kyu;Chang, Yong-Min;Guo, Haiqing
    • Macromolecular research
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    • v.14 no.6
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    • pp.646-653
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    • 2006
  • Highly hydrophilic, uniform, superparamagnetic and nontoxic maltotrionic acid (MA)-coated magnetite nano-particles (MAM) were prepared and characterized by TEM, DLS, XRD and VSM. MA was used to improve the biocompatibility, monodispersity and non-specific intracellular uptake of nanoparticles. Folic acid (FA) was subsequently conjugated to the MAM to preferentially target KB cells (cancer cells) that have folate receptors expressed on their surfaces and to facilitate nanoparticles in their transit across the cell membrane. Finally, fluorescence isothiocyanate (FITC) was added to the nanoparticles to visualize the nanoparticle internalization into KB cells. After the cells were cultured in a media containing the MAM and MAM-folate conjugate (FAMAM), the results of fluorescence and confocal microscopy showed that both types of nanoparticles were internalized into the cells. Nevertheless, the amount of FAMAM uptake was higher than that of MAM. This result indicated that nanoparticles modified with MA and FA could be used to facilitate the nanoparticle uptake to specific KB cells (cancer cells) for molecular imaging.

Serodiagnosis of canine toxoplasmosis by latex agglutination and indirect fluorescent antibody test (Latex 응집반응과 간접형광항체법을 이용한 개 톡소플라즈마병의 혈청학적 진단)

  • Lee, Byung-hoon;Lee, Eung-goo;Suh, Myung-deuk
    • Korean Journal of Veterinary Research
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    • v.32 no.4
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    • pp.641-647
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    • 1992
  • This study was conducted to determine the serum antibodies against toxoplasma in the artificially infected dogs, pet and street dogs by latex agglutination (LA) and indirect fluorescent antibody (IFA) test. LA test was carried out with commercial Toxo-MT kit (Eiken chemical Co.) and IFA test was carried out with rabbit-anti-dog IgG labelled with FITC (Cappel Co.) and toxo-antigen slides prepared in laboratory. The results obtained were as follows ; 1. Antibodies to Toxoplasma gondii in the artificially infected dogs were detected firstly at the Day 8 in IFA and Day 9 in LA test after inoculation. Positive antibody reactions by these tests were declined gradually afterward but maintained up to 12 weeks. 2. In LA test serum antibody titers in 310 test sera were shown as 10 cases(32%) in 1 : 32.5(1.0%) in 1 : 64, 4(1.3%) in 1 : 128 and 2(0.7%) in 1 : 256. In IFA test serum antibody titers 310 test sera were shown as 17 cases(5.5%) in 1 : 64, 8(2.6%) in 1 : 128 and 5(1.6%) in 1 : 256. 3. In the total of 310 sera from pet and street dogs toxoplasma antibody positive rates were 21 cases (6.8%) by LA and 30 cases (9.7%) by IFA test and the positive detection rates between these two groups by LA and IFA test were not significant(p<0.05). 4. In the total of 115 sera from pet dogs toxoplasma antibody positive rates were 12 cases(10.4%) by LA and 15(13.0%) by IFA test. And in the 195 street dogs the positive rates were 9 cases(4.6%) by LA and 15(7.7%) by IFA test. Also, the positive detection rates between these two groups by LA and IFA test were not significant(p<0.05). 5. Agreement of reactivity between LA and IFA test for 310 sera was 91.3% in total of 283 cases consisting of 12 cases(3.9%) of both LA and IFA positive and 271 cases(87.4%) of LA and IFA negative. 6. LA test was almostly equivalent to the IFA test in producibility and proved to be a simple tool for the screening of toxoplasma antibody in laboratory.

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