Résumé
PBS-Tween as a wash solution, prepared with distilled water, is used in ELISA. In areas where schistosomiasis is endemic, however, distilled water is hard to come by. We have modified a WHOLE BLOOD-ELISA test to use coconut water-Tween as a wash solution, because coconut water is easy to come by and cheap in the tropics. We applied the test to whole blood samples from rabbits and humans infected with Schistosoma japonicum. This modified WHOLE BLOOD-ELISA was confirmed to be a rapid, simple, and cost-effective method.
Sujets)
Animaux , Cocos , Test ELISA/méthodes , Humains , Dépistage de masse/méthodes , Lapins , Schistosoma japonicum/enzymologie , Schistosomiase/sang , Tests sérologiques/méthodes , SolutionsRésumé
A rapid and simplified ELISA using whole blood samples of Schistosoma japonicum-infected rabbits was compared with a conventional ELISA. This whole-blood ELISA has advantages. The volume of crude egg antigens, whole blood samples, and conjugates was only 0.05 ml. The incubation time was shortened to 5 minutes. Wells were washed three to five times with PBS-Tween after each procedure. Optical density values were measured in 10 minutes after transfer of 0.1 ml of substrate. Constant temperature was not necessary. The entire procedure took only 20-30 minutes.
Sujets)
Animaux , Antigènes d'helminthe/sang , Test ELISA/méthodes , Dépistage de masse/méthodes , Lapins , Schistosoma japonicum/immunologie , Schistosomiase/diagnostic , Facteurs tempsRésumé
An ELISA technique was developed using samples of Schistosoma japonicum-infected human whole blood based on the conventional ELISA. In this study, the following were demonstrated. 1) Whole blood samples could be used. 2) The volume of whole blood and conjugate could be reduced to 0.05 ml. 3) The incubation time was shortened to 5 minutes. 4) The optical density could be measured at 10 minutes after transferring the substrate and the volume was reduced to 0.1 ml. 5) It did not require a fixed temperature setting. 6) The operation time was as short as 20 to 30 minutes. 7) The optical density values were almost the same as the conventional ELISA and were not influenced by other common intestinal helminthic infections. 8) The observed variations from day to day including effects of sampling in stool examination were negated by the results of this ELISA technique. 9) Based on correlation with stool examination results, criteria can be formulated in which optical density values of 0.3 and above as positive, 0.1 to less than 0.3 as doubtful, and less than 0.1 as negative. Whenever an immunological field survey is necessary, before and after a selective or a mass treatment control program, this WHOLE BLOOD-ELISA, which was shown to be rapid and simple, is recommended.
Sujets)
Adolescent , Adulte , Sujet âgé , Animaux , Antigènes d'helminthe/sang , Enfant , Enfant d'âge préscolaire , Test ELISA/méthodes , Femelle , Humains , Mâle , Dépistage de masse , Adulte d'âge moyen , Schistosoma japonicum/immunologie , Schistosomiase/sangRésumé
The mouse major histocompatibility complex (MHC) class I sequence was detected in all the 8-week-old Schistosoma japonicum recovered from BALB/c (H-2d) and C57BL/6 (H-2b) mice by in situ polymerase chain reaction (in situ PCR). The signals of the mouse class I MHC sequence were observed in the nuclei of the mesenchymal and reproductive cells of 8-week-old S. japonicum. Furthermore, the class I MHC sequence was detected in each DNA extracted from S. japonicum cercariae maintained in BALB/c and C57BL/6 mice by nested PCR. To prove both horizontal and vertical transmission of this sequence in schistosomes, we have used cercariae obtained from parasites maintained in BALB/c mice to infect C57BL/6 and BALB/c mice, and vice versa. The MHC sequences from adult worms were compared to the cercarial MHC and host MHC sequences. Nucleotide sequence comparisons between adult worm DNA, host (H-2d and H-2b mice) DNA and cercarial DNA used for the infection suggested that the sequence of mouse class I MHC was incorporated into schistosome adults and inherited throughout their life-cycle.
Sujets)
Animaux , Séquence nucléotidique , ADN des helminthes/analyse , Modèles animaux de maladie humaine , Transmission de maladie infectieuse/médecine vétérinaire , Transfert horizontal de gène/génétique , Gènes MHC de classe I/génétique , Hétérozygote , Interactions hôte-parasite/génétique , Hybridation in situ , Transmission verticale de maladie infectieuse/médecine vétérinaire , Mâle , Souris , Souris de lignée BALB C/parasitologie , Souris de lignée C57BL/parasitologie , Données de séquences moléculaires , Réaction de polymérisation en chaîne , Schistosoma japonicum/génétique , Schistosomiase artérioveineuse/génétique , Spécificité d'espèceRésumé
Humoral immune responses of IgG, IgM, IgA, IgE and IgG subclass antibodies to Schistosoma japonicum egg antigens were determined by immunoblotting with serum samples from individuals in China with acute (n=24) or chronic (n=35) schistosomiasis. In general, IgM, IgA, and IgE in sera from acute patients exhibited strong binding to antigens but binding was much weaker in chronic cases. Reaction of IgG4 of chronic cases was stronger than that of IgG4 of acute cases. The recognition profile of each antibody isotype in sera was analyzed for 11 major antigen molecules (antigens with apparent molecular weights of 82, 76, 61, 57, 53, 46, 40, 32, 27, 10 and less than 6.5 kDa). Except for the 10 kDa molecule, they were well-recognized by IgA and IgE in sera of acute cases. In other combinations of antibody class and clinical phase, recognition patterns against these molecules differed among individuals. Notably, the 10 kDa molecule was specifically recognized by total IgG and IgG4 in sera from most of the chronic patients, but in sera from only one acute case. This result suggests that the 10 kDa molecule is one of the major target antigens of IgG4 and may be useful as a marker antigen to characterize the clinical phases of S. japonicum infection.