In this study, we are to produce the single chain variable fragment (scFv) antibodies against surface protein of hepatitis B virus (HBV) using antibody phage display technique. Balb/c mice were immunized with preS1 and cDNAs of heavy and light chains of splenic B cells from immunized mice were prepared using RT-PCR. Two cDNAs were linked with (64S) linker DNA under recombination PCR to produce single chain Fv DNA. After digestion of scFv DNA with Sp 1 and Not 1, the digested DNA was ligated into pCANTAB 5E and electroporated into E. coli XL1-Blue to prepare scFv-library. The size of library was 1 * 10' pfu/ml. Phage antibodies (phabs) against preS1 were rescued with M13K07 helper phages, and preS1-binders were selected through 3 times of panning using 96 well microtitre plates. Phage antibody clones were assayed directly for the ability to bind preS1 by ELISA. And then 7 phage antibody clones had high ELISA signals against preS1. Phabs from preS1-specific pMsc-17 had the strongest ELISA signal to preS1. Phabs from pMsc-17 were used for Western blot to preS1 and the results revealed that it was specific to preS1. To prepare the soluble scFv antibody, phabs from pMsc-17 were transfected into non-suppressor E. coli HB2151, and grown under 1 mM IPTG. Soluble scFv antibody was mainly accumulated in the periplasmic space, but small amount of antibody was secreted into culture media.
Animals ; Antibodies ; B-Lymphocytes ; Bacteriophages* ; Blotting, Western ; Cell Surface Display Techniques ; Clone Cells ; Culture Media ; Digestion ; DNA ; DNA, Complementary ; Enzyme-Linked Immunosorbent Assay ; Hepatitis B virus* ; Hepatitis B* ; Hepatitis* ; Isopropyl Thiogalactoside ; Mice* ; Periplasm ; Polymerase Chain Reaction ; Recombination, Genetic ; Single-Chain Antibodies*

Human monoclonal antibodies have considerable potential in the prophylaxis and treatment of viral disease. By cloning human Ig gene segments from the B cells of volunteer into pComb3 phagemid vector, antibody library was created of filamentous phage particles displaying Fab fragments on their surface after being rescued with M13KO7 helper phages. The size of library was 7x10' pfu. Phage antibodies (phabs) were panned against biotinylated preS1 using streptavidine coated Dynabead. The soluble Fab antibodies were prepared from phagemid colonies and assayed directly for the ability to bind preS1 by ELISA. And then 3DW and SGW specific to preS1 which have both heavy and light chain to form Fab fragment, were selected. The soluble Fab antibody from 3DW was expressed highly at the concentration of 0.1 - 1.0 mM of IPTG, and 5 hours postinduction. The soluble antibodies from 3DW and SGW showed their relative affinities of 2x10' M ', and Sx10 M ', respectively, and the specificities to preS1 on ELISA. Our results suggest that antibody phage display library is very useful method to generate the human monoclonal antibody and that the human Fab monoclonal antibodies specific to preS1 selected in this study open the way to treat hepatitis B as a component of passive irnmunotherapeutics.
Antibodies ; Antibodies, Monoclonal ; B-Lymphocytes ; Bacteriophages* ; Clone Cells ; Cloning, Organism ; Enzyme-Linked Immunosorbent Assay ; Genes, Immunoglobulin ; Hepatitis B virus* ; Hepatitis B* ; Hepatitis* ; Humans* ; Immunoglobulin Fab Fragments ; Isopropyl Thiogalactoside ; Streptavidin ; Virus Diseases ; Volunteers

Recently we experienced two cases of cerebral cavernous hemangioma in children at Pediatric Department of Yonsei Medical School. We are reporting these two cases with literature review.
Child* ; Hemangioma, Cavernous, Central Nervous System* ; Humans ; Schools, Medical

No abstract available.
Blotting, Southern*

No abstract available.
Cytogenetics* ; Korea* ; Oncogenes* ; Stomach Neoplasms*

OBJECTIVE: To investigate the conduction velocity of sympathetic skin response(SSR) in normal adults. METHOD: The latency of SSR was measured in 41 normal healthy subjects by the simultaneous recordings from three sites of the hand. And we also measured the distance and conduction time between the recording sites of the hand. The conduction velocity of SSR was calculated by dividing the distance by conduction time. RESULTS: The SSR was obtainable in all subjects from three sites of the hand. The mean latencies of SSR recorded from wrist, midpalm and index finger were 1.29, 1.40 and 1.54 seconds respectively. And the mean latency showed a significant increase from wrist to index finger(p<0.05). The conduction velocity of the SSR from wrist to index was 0.57 m/sec, and segmental conduction velocities from wrist to palm and palm to index were 0.62 and 0.66 m/sec respectively. The conduction velocity of SSR in the distal segment was slightly faster than in the proximal segment with no statistical significance. CONCLUSION: The conduction velocity of SSR by the simultaneous recordings at two or more sites of the hand can be easily obtained and offers a useful parameter along with the amplitude and latency of SSR.
Adult* ; Fingers ; Hand ; Humans ; Skin* ; Wrist

The double stapling technique has become an established reconstruction method for patients undergoing low anterior resection. We have used a modification of the conventional technique in which the lower rectal segment is closed with a linear stapler (TA-55) and the anastomosis is performed by using the circular EEA(CEEA) instrument across the linear staple line of the double stapling technique. The aim of this study was to evaluate the prophylactic effect of a loop ileostomy preventing anastomotic leakage. Stapled colorectal anastomosis and stapled coloanal anastomosis in 60 patients forms the basis for the report. The sixty patients were treated by using the double stapling technique either with or without a loop ileostomy. This review presents the advantages and disadvantages of a loop ileostomy for coloanal anastomosis. Postoperative anastomotic leakage in the double stapling technique group occurred in 5 (10.6%)cases of the total 31 cases while in the double stapling technique with loop ileostomy group, it allowed 1(3.4%) of the total 29 cases. This study suggests that the double stapling technique with a loop ileostomy is more effective than the double stapling technique without a loop ileostomy in preventing anastomotic leakage. The addition of a loop ileostomy to protect the low anastomosis might also be expected to influence anastomotic healing.
Anastomotic Leak ; Humans ; Ileostomy ; Rectal Neoplasms*

Duchenne muscular dystrophy(DMD) is an X-linked recessive disease, caused by the mutation of dystrophin gene at Xp21. The dystrophin produced by this gene is therefore absent on the membrane of muscular fiber in the patie nts with DMD. Recently, it is known that the dystrophin has also been located on the myoepithelial layer of sweat gland in the mice. We studied the sympathetic skin response(SSR) in a group of DMD patients and a control group to evaluate the function sympathetic nerve and sweat gland in DMD patients. Significant prolongation of latency of SSR in the palm and sole was noted in the group of DMD patients compared to the control group. However, there was no significant difference in the amplitude of SSR between two group. In the patient group, the rise in latency of SSR was closely correlated with the duration of symptoms and weakly associated with the stage of the illness. Therefore the latency of SSR may be a useful index in assessing the function of sympathetic nerve and sweat gland in DMD patients. These results could be a consequence of a lack of dvstrophin at myoepithelium of sweat gland in DMD patients.
Animals ; Dystrophin ; Humans ; Membranes ; Mice ; Muscular Dystrophy, Duchenne* ; Skin* ; Sweat Glands