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 1,323 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Soybean mosaic virus (SMV) is an important disease in soybean and is widely distributed in northern of Iran. SMV transmitted by soybean seed and detection of it is very important for disease management. In this study, several detection methods including DAS-ELISA, indirect-ELISA, tissue-print immunoassay (TPIA) and Dot immunobinding assay (DIBA) were optimized and compared with each other to identify the virus, using polyclonal antibody. For TPIA, nitrocellulose membrane was used to imprint fresh sections of healthy and infected plant materials, and for DIBA 10 microl of extracts was doted onto nitrocellulose membranes. Both membranes were incubated 1 hour in blocking buffer, and then incubated 2 h in 1:1000 dilution of IgG-conjugate. After incubation the membranes were washed three times with PBS-T buffer for 15 min. Then the membranes were incubated in substrate solution containing NBT/BCIP. After some minutes prints or blots of infected tissues turned dark violet, whereas prints or blots of healthy ones did not show any color changes. In some cases, substrate solution was Fast red, containing 0.2M Tris-HCl buffer and 2mM MgCl2, pH = 7.8, producing red color in infected prints or blots. Both methods are simple and TPIA is rapidly and easily applicable in the field. However, TPIA had some advantages over the others. TPIA is time-saving as there is no need for conventional sap extraction and also nitrocellulose membranes used for printing can be used in the field and stored for a long time or transported to another laboratory for process. These two methods can be used routinely for detection of SMV in many samples.
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PMID:Rapid diagnosis of soybean mosaic virus N soybean by tissue-print immunoassay and DIBA in comparison to other serological methods. 1739 Aug 80

Field experiments are usually necessary for analyzing the efficiency of control methods, the resistance of varieties and many other virological studies. Such experiments generally include a large number of samples to be tested serologically. DAS-ELISA is a very accurate technique that has been widely used for identifying viral infections. For large numbers of plant samples, it takes a long time for the sample preparation, plate washing and other procedures. In this study, the efficiency of a simple and laborsaving TPIA (Tissue Print Immunoassay) method was evaluated for the identification of two important aphid-transmitted viruses (CMV and PVY) of tobacco fields in comparison with DAS-ELISA as the standard method. The leaf samples were collected from the fields of three commercial tobacco types (flue-cured, burley and oriental). Each sample was divided to two parts and each part was examined by one of the methods. DAS-ELISA was done based on the method described by Clark and Adams (1977). In TPIA, small parts of the leaf samples were rolled and then cut by a sterile sharp blade. The cut surface was gently printed on 1 cm2 blocks drawn on a nitrocellulose paper. Air dried paper was located first in 1% BSA for blocking the empty sites on paper, then in the buffer containing AP-conjugated polyclonal antibody for 3 h and finally in NBT-BCIP solution for color development. Between these stages, the paper was washed thoroughly (three times) by shaking in fresh washing buffer. The results of each sample were recorded and compared with those of DAS-ELISA. By considering DAS-ELISA as the reference method, the sensitivity of TPIA for the detection of PVY and CMV was 96.1% and 92.7%, respectively. The positive results by TPIA which were not detected positive by DAS-ELISA were regarded as false positive. These were 8 (out of 316 tested samples) for CMV and 6 (out of 204 samples) for PVY. Although the results of TPIA were not completely consistent with DAS-ELISA, it seems that this method can be used for some general studies. The most important advantages of this method were that it didn't need sample extraction and was done using only one antibody which was the conjugated antibody of each virus. This method gives more rapid results (within a day) in comparison with DAS-ELISA that needs more time.
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PMID:Evaluating the efficiency of a TPIA method for the detection of two tobacco viruses. 1739 Aug 82