We have applied an innovative commercial test system (the GeneGen Major Food Pathogens [MFP] Detection Kit) to detect four of the more important pathogenic microorganisms in food, Salmonella spp., Escherichia coli O157, Listeria monocytogenes and Campylobacter spp. by using multiplex polymerase chain reaction (PCR) combined with a shortened, simplified hybridization protocol on test strips. Testing 129 strains representing 23 different genera yielded no false‐positive or false‐negative results, and the limit of detection was 100 genome copies of each organism per multiplex PCR using purified genomic DNA. The kit was used for both analyses of chicken samples artificially contaminated with different numbers of the pathogens and of naturally contaminated samples. The results were compared with those obtained with standard microbiological techniques. Both methods yielded the same results in spiking experiments, detecting 1–10 cfu/25 g of the respective pathogen added. It was possible to verify the presence of all four pathogens simultaneously in one assay. The GeneGen MFP Detection Kit saves at least 2–4 days compared with standard methods. The protocol is easy to perform, and there is no need for any additional costly and sophisticated equipment, except for a thermocycler, because hybridization is carried out at ambient temperature and the results can be evaluated with the unaided eye. This enables the assay to be performed in relatively modestly equipped laboratories for both DNA‐based screening of food samples and/or confirmation of suspect colonies.
A chimeric HIV-2/HIV-1 envelope sequence containing an immunodominant region of HIV-2 gp36 and the corresponding region of HIV-1 gp41 was constructed and overexpressed in Escherichia coli. The recombinant product (rp21/18) was purified and applied in a novel antibody-screening assay. Characteristics in the design of this new principle are as follows: (1) the overall assay time is about 30 min; (2) the assay procedure includes three manipulation steps; and (3) the test shows a reliable performance with respect to sensitivity and specificity. The diluted serum sample and the protein G-horseradish peroxidase conjugate are added simultaneously into a coated (hybrid antigen HIV-1/2) and blocked microtiter plate well. The in-batch incubation of serum sample with protein G-horseradish peroxidase saves two manipulation steps that are normally necessary in the five-step procedure of a classical ELISA. AIBS was evaluated with commercially available seroconversion panels and with random negative serum samples from a blood bank. Seroconversion results demonstrated that AIBS has equivalent sensitivity to ELISAs and the third generation assays. The specificity was determined on a total blood donor population of 5012 (Red Cross Vienna, Austria). The repeat reactive rates for donor population were 0.02%. AIBS represents a general immunometric system (not only HIV antibodies). The entire assay procedure of AIBS evaluated for HIV-1/2 screening, including result reporting, can be performed automatically by several commercially available systems. Depending on these systems AIBS is potentially useful in laboratories or blood banks that have both high- and low-volume testing.
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