• BMB Reports
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• v.38 no.6
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• pp.703-708
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• 2005

We cloned and expressed human pyridoxal-5'-phosphate (PLP) phosphatase, the coenzymatically active form of vitamin $B_6$, in Escherichia coli using pET15b vector. Monoclonal antibodies (mAb) were generated against purified human brain PLP phosphatase in mice, and four antibodies recognizing different epitopes were obtained, one of which inhibited PLP phosphatase. The binding affinities of these four mAbs to PLP phosphatase, as determined using biosensor technology, showed that they had similar binding affinities. Using the anti-PLP phosphatase antibodies as probes, we investigated their cross-reactivities in various mammalian and human tissues and cell lines. The immunoreactive bands obtained on Western blots had molecular masses of ca. 33 kDa. Similarly fractionated extracts of several mammalian cell lines all produced a single band of molecular mass 33 kDa. We believe that these PLP phosphatase mAbs could be used as valuable immunodiagnostic reagents for the detection, identification, and characterization of various neurological diseases related to vitamin $B_6$ abnormalities.

• BMB Reports
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• v.39 no.2
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• pp.208-215
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• 2006

The human ribosomal protein S3 (rpS3) was expressed in E. coli using the pET-I5b vector and the monoclonal antibodies (mAbs) were produced and characterized. A total of five hybridoma cell lines were established and the antibodies recognized a single band of molecular weight of 33 kDa on immunoblot with purified rpS3. When the purified rpS3 was incubated with the mAbs, the UV endonuclease activity of rpS3 was inhibited up to a maximum of 49%. The binding affinity of mAbs to rpS3 determined by using a biosensor technology showed that they have similar binding affinities. Using the anti-rpS3 antibodies as probes, we investigated the cross-reactivities of various other mammalian brain tissues and cell lines, including human. The immunoreactive bands on Western blots appeared to be the same molecular mass of 33 kDa in all animal species tested. They also appear to be extensively cross-reactive among different organs in rat. These results demonstrated that only one type of immunologically similar rpS3 protein is present in all of the mammalian brain tissues including human. Furthermore, these antibodies were successfully applied in immunohistochemistry in order to detect rpS3 in the gerbil brain tissues. Among the various regions in the brain tissues, the rpS3 positive neurons were predominantly observed in the ependymal cells, hippocampus and substantia nigra pars compacta. The different distributions of rpS3 in brain tissues reply that rpS3 protein may play an important second function in the neuronal cells.

• Journal of Food Hygiene and Safety
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• v.31 no.6
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• pp.445-450
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• 2016

Thermal-stable soluble proteins (TSSP) in livestock products has been recently reported. Therefore, the development of antibodies and immunoassay using a TSSP is useful because the presence of TSSP can be measured on processed food. In this study, the existence of TSSPs in pork fat and meat was confirmed and their antigenicity was investigated. The extracts from pork fat and meat by heating method were analyzed by SDS-PAGE with 5% stacking and 12% separating gels. The protein profiles from the raw pork fat and meat extracts (major band ranged 25 to 100 kDa) without cooking and heating treatments were significantly different compared to those from cooked and heated pork fat and meat extracts (several major bands > 100 kDa and < 30 kDa). This meant that non thermal-stable soluble proteins ranged from 25 to 100 kDa may be denaturated to insoluble proteins by cooking and heating treatments, and TSSPs were in pork fat and meat at kept their properties. The confirmed TSSPs were used as an immunogen to investigate their antigenicity. Eight mice (5 mice for pork fat and 3 mice for pork meat) were separately immunized with the TSSPs of pork fat and meat, and the anti-sera obtained from the immunized mice showed high titer values. Polyclonal antibodies against each target protein showed the specific reaction to pork fat and meat, individually. These indicated that TSSP could be used as an immunogen to produce antibodies such as monoclonal and polyclonal antibodies. In addition, antibodies specific to TSSP from pork fat and meat may be used as a bio-receptor in immunoassays for the identification of fraudulent adulteration with pork fat and meat in livestock products.

• Clinical and Experimental Pediatrics
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• v.48 no.5
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• pp.506-511
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• 2005

Purpose : Streptococcus pneumoniae serotype 6B and 6A are important pathogens in pneumococcal infections. It is commonly assumed that the 6B vaccines elicit antibodies cross-reacting with the 6A serotype and the cross-reactive antibodies protect against infections of 6A. To examine this assumption, we measured the opsonophagocytic capacity to serotype 6A and 6B in adults. Methods : Twenty-four adults were immunized with pneumococcal PS vaccine that contains 6B PS. Their preimmune and postimmune sera were studied for the capacity to opsonize 6B and 6A serotypes with opsonophagocytic killing assay. Results : Opsonization titers to 6B were significantly higher than those to 6A in preimmune and postimmune sera. Because significant increasesof opsonization titers were observed in adults with polysaccharide vaccines for 6A(cross-reactive) serotype as well as for 6B(vaccine) serotype, 6B PS in vaccine elicited cross-protective antibodies to 6A, but not in all cases. One adult did not have detectable levels of opsonization titers to 6A after immunization. Conclusion : Although 6B PS in pneumococcal PS vaccine elicits antibodies cross-reacting with 6A serotype in some adults, it may not occur always. This study should be extended to other age groups such as children and elderly people. The presence of the cross-protection should be directly determined in clinical trials of the pneumococcal vaccines as well as during the postlicensure monitoring surveys by serotyping the clinical isolates of pneumococci.

• Archives of Pharmacal Research
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• v.27 no.9
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• pp.961-967
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• 2004

The development of an antibody labeling method with $^{99m}$Tc is important for cancer imaging. Most bifunctional chelate methods for $^{99m}$Tc labeling of antibody incorporate a $^{99m}$Tc chelator through a linkage to lysine residue. In the present study, a novel site-specific $^{99m}$Tc labeling method at carbohydrate side chain in the Fc region of 2 antibodies (T101 and rabbit anti-human serum albumin antibody (RPAb)) using dihydrazinophthalazine (DHZ) which has 2 hydrazino groups was developed. The antibodies were oxidized with sodium periodate to pro-duce aldehyde on the Fc region. Then, one hydrazine group of DHZ was conjugated with an aldehyde group of antibody through the formation of a hydrazone. The other hydrazine group was used for labeling with $^{99m}$Tc. The number of conjugated DHZ was 1.7 per antibody. $^{99m}$Tc labeling efficiency was 46-85％ for T101 and 67∼87％ for RPAb. Indirect labeling with DHZ conjugated antibodies showed higher stability than direct labeling with reduced antibodies. High immunoreactivities were conserved for both indirectly and directly labeled antibodies. A biodistribution study found high blood activity related to directly labeled T1 01 at early time point as well as low liver activity due to indirectly labeled T101 at later time point. However, these findings do not affect practical use. No significantly different biodistribution was observed in the other organs. The research concluded that DHZ can be used as a site-specific bifunctional chelating agent for labeling antibody with $^{99m}$Tc. Moreover, $^{99m}$Tc labeled antibody via DHZ was found to have excellent chemical and biological properties for nuclear medicine imaging.edicine imaging.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.48 no.2
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• pp.158-167
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• 2015

A lateral flow immunoassay kit based on antigen-antibody interactions was developed to detect residues of beta-lactams, quinolones, tetracyclines, and sulfonamides in farmed fish. Group-specific antibodies showing cross-reactivity with other antibiotics in the same group were produced in rabbits. The rabbits were immunized eight times to obtain the maximum titers. Antibodies were extracted from the antisera collected from the immunized rabbits and produced group-specific reactions with antibiotics from the four groups. A kit was prepared that optimize conditions for the antigen-antibody reaction, using colloidal gold conjugated antibodies, and was designed to detect the four groups of antibiotics simultaneously. The kit enabled the detection of antibiotics in the four groups at below maximum residue limits (MRLs), which were $200{\mu}g/kg$ for tetracyclines, $100{\mu}g/kg$ for sulfonamides, $50{\mu}g/kg$ for beta-lactams, and $100{\mu}g/kg$ for quinolones. The cross-reactivity of the antibodies ranged from 10-80% for the sulfonamides, 20-100% for tetracyclines, 38-100% for quinolones, and 20-100% for the beta-lactams, confirming that the antibodies were group specific. The test kit was used 30 times to examine spiked antibiotics at the limits of detection (LODs) and all produced positive results, indicating high sensitivity. The LODs for the assay ranged from 4-20 ng/mL for beta-lactams, 25-50 ng/mL for sulfonamides, 20-100 ng/mL for tetracyclines, and 30-80 ng/mL for quinolones, and there were no false negative reactions at above these LODs. In addition, all of the LODs of the developed kit were correlated with high-performance liquid chromatography (HPLC) data. Our lateral flow immunoassay kit can simultaneously detect antibiotic residues from a large number of fish samples rapidly, strengthening the safety of domestic farmed and imported fish.

• Korean Journal of Agricultural Science
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• v.14 no.2
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• pp.401-408
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• 1987

Nine monoclonal antibodies directed against infectious bovine rhinotracheitis virus (IBRV) were prepared by using cell hybridization technique, and the biological properties of the antibodies were investigated by means of immunofluorescence, serum neutralization, and electrophoretic analysis. Eight of 9 monoclonal antibodies reacted specifically with the antigenic constituents of IBRV, infectious laryngotracheitis virus, Marek's disease virus, turkey herpesvirus, hog cholera virus, porcine parvovirus and transmissible gastroenteritis virus. However, the remaining one, 26-2 clone, was found to be cross-reactive with pseudorabies virus. Two monoclonal antibodies, 7-C-2 and 12-A-2, which had neutralizing activity, were reactive with the molecular weights of 72 kilo daltons (72K) and 125K of IBRV proteins electrophoretically separated, respectively. The monoclonal antibody, 3-H-3, which is corresponding to 94K of IBRV proteins, revealed no neutralizing activity. The cross-reactive monoclonal antibody, 26-2, was proved by electrophoretical analysis to be reactive with 100K of IBRV proteins and 40K of pseudorabies virus.

• Food Science and Biotechnology
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• v.16 no.4
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• pp.515-519
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• 2007

Rapid immunochromatographic assay (ICA) kits were developed using flagella-specific monoclonal antibodies (MAbs) and rabbit polyclonal antibodies for screening Listeria spp. in food. The establishment of different formats, MAb 2B1 as capture antibody and MAb 7A3 or rabbit polyclonal antibodies as detector antibody, was compared. The 2 formats of the ICA kit were shown to have specific reactions with Listeria and no cross-reactivity with any of the non-Listeria including Escherichia coli O157:H7 and Salmonella enteritidis. The detection limits of the ICA kit using the combination of gold-labeled MAb 7A3 and MAb 2B1 showed $1{\times}10^5$ and $1{\times}10^6\;CFU/0.1\;mL$ at 22 and $30^{\circ}C$, respectively. The other format of the ICA kit using the combination of gold-labeled rabbit polyclonal antibodies and MAb 2B1 showed $1{\times}10^6\;CFU/0.1\;mL$ at $22^{\circ}C$ but weak signal at 30 culture. The format utilizing MAb was more sensitive than the one using polyclonal antibodies for capture antibody. Samples contaminated with L. monocytogenes 4b culture (9-10, 5-6, and 1-2 CFU/mL) on pork and pasteurized milk were confirmed as positive results. Current data suggests that this ICA kit is a rapid, simple and effective tool to screen for Listeria spp. in food.

• Parasites, Hosts and Diseases
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• v.33 no.3
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• pp.187-194
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• 1995

The present study aims to observe changing patterns of serum antibody to hleumuvstis calinii in normal rats of different ages and in immunosuppressed rats. The serum IgG antibody was observed by immunoblotting with crude antigen of f carinii which were purified from the lungs of infected rats. The crude antigens separated in SDS-PAGE resolved more than 20 protein bands from 20 to 200 kDa. Of them,40-45, 50-55, 116 and 200 kDa bands were major antigens of R cori.nii. Most of the normal rats of up to 4 weeks had the antibodies reacting the 4 bands, but none of 8-week-old rats revealed the specific antibody. After the rats grew for 40 weeks, all were found to have the antibody in their serum. Same pattern of serum antibody level by age was found in ELISA. When immunosuppressed rats became heavily infected, the antibody in their serum decreased distinctively. The present results suggest that antibodies in normal newborn rats are transferred from their mother and lowered up to 8 weeks. Thereafter, the levels of the antibodies begin to increase by natural exposure to R cnrinii. It was also confirmed that the intensity of P cnrinii infection is inversely related with levels of serum antibodies.

• Journal of Animal Science and Technology
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• v.51 no.3
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• pp.231-240
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• 2009

Sixteen ruminally cannulated Korean native steers (Hanwoo; $626.2\pm47.72$ kg) were used to investigate the effects of polyclonal antibodies against abdominal (AAb) and subcutaneous adipocyte membrane proteins (SAb) on ruminal fermentation patterns and blood metabolites. The body weight (BW) of Hanwoo was decreased 2-weeks after AAb and SAb injection, BW reduction was also observed in control and non-immunized serum groups, indicating that stress induced by other factors (e.g. blood sampling etc.) rather than antibodies injection may affect the BW reduction. Antibodies treatment did not affect (P > 0.05) rumen pH, volatile fatty acids and ammonia-N concentration. The ranges were similar with typical ranges of those in Hanwoo. Compared with control, blood urea N concentration was decreased in AAb group and increased (P < 0.05) in SAb group before antibodies treatment. However, none of the groups were significantly (P > 0.05) affected at 2- or 4-weeks after the treatment. Concentration of plasma glucose in the non-immunized serum group was significantly higher (P < 0.05) than the other groups at 0-week after treatment. However, antibodies treatment did not affect the concentration of plasma glucose. Concentration of plasma triglyceride showed no difference (P > 0.05) between the groups and ranged from 11.4 to 19.9 mg/dl, which is the perfect range of plasma triglyceride of Hanwoo fed concentrate based diets. In conclusion, these results may indicate that the present AAb and SAb have safety in nutritional physiological metabolism in Hanwoo. Further study on in vivo fat reduction of the antibodies against abdominal and subcutaneous adipocytes PMPs of Hanwoo is required for inedible fat-reduced high quality beef production.