Functional heavy-chain antibodies in Camelidae.
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Animals↗
Biomedical subjects
V K Nguyen
Publications and source records attributed to V K Nguyen.
Camel heavy-chain antibodies: diverse germline V(H)H and specific mechanisms enlarge the antigen-binding repertoire.
The antigen-binding site of the camel heavy-chain antibodies devoid of light chain consists of a single variable domain (V(H)H) that obviously lacks the V(H)-V(L) combinatorial diversity. To evaluate the extent of the V(H)H antigen-binding repertoire, a germline database was constructed from PCR-amplified V(H)H/V(H) segments of a single specimen of Camelus dromedarius. A total of 33 V(H)H and 39 V()H unique sequences were identified, encoded by 42 and 50 different genes, respectively. Sequence comparison indicates that the V(H)Hs evolved within the V(H) subgroup III. Nevertheless, the V(H)H germline segments are highly diverse, leading to a broad structural repertoire of the antigen-binding loops. Seven V(H)H subfamilies were recognized, of which five were confirmed to be expressed in vivo. Comparison of germline and cDNA sequences demonstrates that the rearranged V(H)Hs are extensively diversified by somatic mutation processes, leading to an additional hypervariable region and a high incidence of nucleotide insertions or deletions. These diversification processes are driven by hypermutation and recombination hotspots embedded in the V(H)H germline genes at the regions affecting the structure of the antigen-binding loops.
Amino Acid Sequence↗
Protection of immunoreactivity of dry immobilized proteins on microtitration plates in ELISA: application for detection of autoantibodies in myasthenia gravis.
We show the ability of the BSA-trehalose film to convert normally fragile proteins such as mouse monoclonal antibody to the Alzheimer precursor protein A4 (APP695) and cell line TE671 acetylcholine receptor (AChRTE671) into a stable reagent, after its immobilization on microtitration plates. The remarkable property of the dry immobilized proteins are their stability under prolonged exposure to temperatures as high as 50 degrees C. Using the AChRTE671, the proposed method was applied for the measurement of anti-AChR autoantibodies in Myasthenia gravis by means of an enzyme-linked immunosorbent assay (ELISA). The test was shown to be specific and able to detect anti-AChR autoantibodies at concentrations as low as 3 nM. Using the same AchRTE671 as antigen, the results of examination of 34 serum samples for detection of anti-AChR autoantibodies by ELISA were compared with those of the conventional radioimmunoprecipitation assay (RIA). It was concluded that ELISA is another useful method for the diagnosis of M. gravis. The ELISA method offers a rapid, simple, safe and inexpensive means for mass screening of M. gravis.
Autoantibodies↗
'LABNOTE', a laboratory notebook system designed for academic genomics groups.
We have developed a relational laboratory database system, adapted to the daily book-keeping needs of laboratories that must keep track of information acquired on hundreds or thousands of clones in an effective and user-friendly fashion. Data, whether final or related to experiments in progress, can be accessed in many different ways, e.g. by clone name, by gene, by experiment or through DNA sequence. Updating, import and export of results is made easier by specially developed tools. This system, in network version, serves several groups in our Institute and (over the Internet) elsewhere, and is instrumental in collaborative studies based on expression profiling. It can be used in many similar situations involving progressiveaccumulation of information on sets of clones or related objects.
Database Management Systems↗
Loss of splice consensus signal is responsible for the removal of the entire C(H)1 domain of the functional camel IGG2A heavy-chain antibodies.
The molecular basis for the absence of the C(H)1 domain in naturally occurring heavy-chain antibodies of the camelids was assessed by determining the entire Camelus dromedarius gamma2a heavy-chain constant gene. The organization of the camel gamma2a constant heavy-chain gene obtained from a liver genomic library appears to be typical of all other mammalian gamma genes sequenced to date. It contains the switch, CH1, hinge, CH2, CH3, M1 and M2 exons. In contrast to the case in mouse and human heavy chain diseases, the camel gamma2a gene shows no major structural defect, and its equivalent CHI exon is intact. However, sequence analysis has revealed that the splicing site, immediately after the CH1 exon, is defective due to point mutations, especially the G(+1) to A(+1) transversion seems to be detrimental. It is concluded that the loss of the splice consensus signal is responsible for the removal of the entire CH1 domain in camel gamma2a heavy-chain immunoglobulins. Additionally, a closer analysis of the hinge exon suggests the possible involvement of transposons in the genetic variation of mammalian Cgamma hinges.
Amino Acid Sequence↗
[Reactions of cells of nasal and sinusoidal mucosa of goats and sheep naturally infected by Oestrus ovis Linné 1758 (Diptera: Oestridae)].
OEstrosis is a very common myiasis of sheep and goats in mediterranean and tropical countries. Goats are suitable hosts for OEstrus ovis but the parasitic burden remain lower than in sheep. Cellular responses (mucous and serous mast cells, eosinophis and globules leucocytes) were measured in 30 infected and 30 non infected sheep and in 23 infected and 24 non infected goats. The presence of OEstrus ovis larvae led to an important inflammatory response in sheep and in goats as well. Furthermore, the intensity of the cellular response correlated with the larva burden, specially with globules leucocytes and eosinophils. Nevertheless, huge differences occurred between sheep and goats responses even in similar larval burden range. Infected sheep showed larger counts of mucous mast cells than goats, the differences were smaller in serous mast cells and eosinophils and no difference was detected in globules leucocytes (intraepithelial mast cells) counts between the two hosts species. These results were compared with those obtained in gastro-intestinal strongyles infections.
Animals↗
The specific variable domain of camel heavy-chain antibodies is encoded in the germline.
The variable domains of the functional heavy-chain antibodies (VHHs) discovered in camels are related to the human VH subgroup III. They are nevertheless clearly distinguishable from the VHs of conventional four-chain immunoglobulins by the presence of important amino acid substitutions, located in the solvent-exposed surface normally covered by the variable domain of the light chain. The analysis of an unrearranged dromedary DNA library revealed that the specific VHH gene with its characteristic amino acid substitutions is encoded in the germline. Therefore, it is concluded that the VHHs do not arise through an ontogenic process of somatic hypermutation. The presence of putative DNA recombination signals that are more prevalent in the camel VHH, compared to the VH germline gene, might play a role in the formation and efficient expansion of the VHH repertoire.
Amino Acid Sequence↗
[Mast cells and eosinophils of the respiratory mucosa of sheep with Oestrus ovis (Linné, 1761) infection].
Mast cells and eosinophils have been identified by differential stainings and counted in mucous membrane of nasal septum, turbinates and sinus of 77 ewes naturally infected with Oestrus ovis. Results have been compared with those of nine parasite free lambs. Anova tests indicate significant differences between infected and parasite-free sheep for the cell numbers and their distribution among the septum, the turbinates and the sinus and according to their position in mucous membrane, interglandular chorion of sub-mucosa. In infected sheep, the mean number of mast cells is twice the number present in parasite free animals. The burdens of eosinophils are multiplied by 17 for the septum, 29 for the turbinates and 58 for the sinus. The hypothesis of the development of an hypersensitivity phenomenon in ovine oestrosis is sustained by these results.
Animals↗
Evaluation of an enzyme-linked immunosorbent assay for detection of antibodies to bovine leukemia virus in serum and milk.
The results of examination of 375 bovine serum and 150 bovine milk samples for detection of bovine leukemia virus infection by the immunodiffusion technique were compared with those of an enzyme-linked immunosorbent assay (ELISA). It was concluded that the ELISA is another useful method for the detection of antibodies to bovine leukemia virus in serum and milk. The ELISA provides a quantitative result and has the advantage of being more sensitive and less time-consuming than the conventional immunodiffusion technique.
Animals↗
Immobilized enzyme electrode for creatinine determination in serum.
An immobilized enzyme electrode for continuous creatinine determination in blood serum is described. The enzymes creatinine amidohydrolase, creatine amidinohydrolase, and sarcosine oxidase are coimmobilized to the surface of the polypropylene membrane of a Clark-type electrode responsive to oxygen. The immobilized enzymes catalyze the decomposition of creatinine with the consumption of oxygen and thus permit the creatinine measurement. The whole assay takes less than 1 min. Effects of pH and temperature on electrode response are also described. The proposed technique offers a rapid, simple, and inexpensive means to determine creatinine in blood serum within the normal and abnormal ranges. The repeatability of the creatine determination in serum is 2.5% (relative standard deviation), and the detection limit is 3 x 10(-6) mol L-1. The results obtained by this method were compared to those obtained with the Technicon AutoAnalyzer SMAC system based on the Jaffé reaction; the correlation factor between the two methods was found to be r = 0.9997.
Amidohydrolases↗
Enzyme immunoassay (ELISA) for detection of Clostridium difficile toxin B in specimens of faeces.
Antisera against Clostridium difficile toxin B were prepared in sheep and rabbit and were used in indirect and sandwich enzyme-linked immunosorbent assays (ELISA) for the detection of toxin B. Polyvinyl chloride and polystyrene microtitration plates were tested as solid phases for the assay. Both assays had a lower limit of detection for toxin B of 1 ng/ml. They were used to detect the presence of toxin B in 210 human faecal specimens and also in the culture supernatant fluids of C. difficile strains isolated from the faecal samples. There was a close correlation between the results of sandwich ELISA and those of cytotoxicity tests and isolation of C. difficile. Our sandwich ELISA method seems to be useful as a presumptive test for detection of C. difficile toxin B.
Bacterial Proteins↗
Clostridium difficile toxin B: characterization and sequence of three peptides.
The cytotoxin, also named toxin B, was isolated from a toxigenic strain of Clostridium difficile, purified to homogeneity and partially characterized. The purification procedure included ultrafiltration followed by anion-exchange chromatography. We noticed that a non-specific nucleic material eluted with the protein during the purification. The presence of these nucleic acids appeared to be important for the toxic activity of the protein. Some characteristics of the cytotoxin were examined, especially the amino acid composition and the sequence of three tryptic fragments.
Amino Acid Sequence↗
Fast protein purification of Clostridium difficile cytotoxin.
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Bacterial Proteins↗
Double-blind studies with mefloquine alone and in combination with sulfadoxine-pyrimethamine in 120 adults and 120 children with falciparum malaria in Vietnam.
In 120 adult Vietnamese patients with uncomplicated falciparum malaria the efficacy of, and tolerance to, mefloquine (M) vs the combination of mefloquine + sulfadoxine + pyrimethamine (MSP) was studied in a double-blind, randomized comparative trial with chloroquine. Also, a double-blind dose finding study of MSP was performed in 120 Vietnamese children with uncomplicated falciparum malaria. In the adults the mean parasite clearance time with M was 3.8 d and with MSP 3.6 d. Defervescence occurred in 2.9 and 3.0 d respectively for M and MSP. There was a 36.8% resistance rate in 38 patients treated with chloroquine. 96% of the children were sensitive or showed a delayed RI response. The lowest dose of MSP (10 mg/kg M + 20 mg/kg S + 1.0 mg/kg P, 1 tablet Fansimef) was as effective as 1.5-2x this dose in children weighing 23-30 kg. Side effects were mild, except for vomiting which required alternative therapy in 4 patients.
Adolescent↗
[The efficacy of and tolerance for fansimef in the treatment of tropical malaria in the south of the Socialist Republic of Vietnam].
The efficacy of and tolerance to fancimef in 49 patients was compared to the efficacy of and tolerance to quinine in combination with fansidar in 33 patients with moderate P. falciparum malaria. A good tolerance to and a high efficacy of fancimef have been shown, which was manifested in rapid fever arrest, disappearance of parasitemia, absence of the disease relapses, which were observed in 2 patients on quinine combined with fansidar. The advantage of fansimef is one-time administration of the total course dose.
Acute Disease↗
[Liposarcoma of the mediastinum].
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Female↗
[Ca++ ion transport blockers as reversants of the drug resistance of malarial parasites. 1. The effect of verapamil on the resistance to chloroquine in vivo of Plasmodium berghei and in vitro of Plasmodium falciparum].
The reversing action of verapamil on the effect of chloroquine was found in in vivo experiments by using a model P. berghei resistant to chloroquine, an LNK65 isolate having a naturally lower resistance to the agent, and its polyresistant strain with the acquired resistance to chloroquine and fansidar, as well as by employing the chlorine-resistant P. falciparum isolates from the south of the Socialist Republic of Vietnam. The magnitude of this effect was related to the dose of verapamil, the frequency of administration of a combination of the agents in vivo, while that was associated to the concentration of verapamil and the level of isolate resistance to chloroquine in vitro which was the most pronounced. Taking into account the dose-dependent effect of verapamil, it can be suggested that increasing its concentration in combination with chloroquine can provide a more marked reversing action with lower chloroquine concentrations. The parameters accepted by the authors in evaluating the combined effect enable the effect of the verapamil/chloroquine concentration to be regarded as potentiation.
Animals↗