A novel non-radioactive assay for HIV-RT(RdDp)based on pyrosequencing for high-throughput drug screening

Current in vitro assays for the activity of HIV-RT(reverse transcriptase)require radio-labeled or chemically modified nucleotides to detect reaction products.However,these assays are inherently end-point measurements and labor intensive.Here we describe a novel non-radioactive assay based on the principle of pyrosequencing coupledenzyme system to monitor the activity of HIV-RT by indirectly measuring the release of pyrophosphate(PPi),which is generated during nascent strand synthesis.The results show that our assay could monitor HIV-RT activity with high sensitivity and is suitable for rapid highthroughput drug screening targeting anti-HIV therapies due to its high speed and convenience.Moreover,this assay can be used to measure primase activity in an easy and sensitive manner,which suggests that this novel approach could be wildly used to analyze the activity of PPi-generated and ATP-free enzyme reactions.

Evaluation of a sulfonated DNA probe (sulfoprobe) for diagnosis of falciparum malaria

In this study,the DNA probe pPF14 was nonradioactively labelled by sulfo-modifica-tion,and used in a dot blot hybridization assay for detection of P.falciparum.The resultsshowed that the sulfoprobe successfully detected 25pg purified DNA and 0.001% parasitemia ofcultured P.falciparum,and did not react with human leukocyte DNA.In the study of 179clinical blood samples of malaria patients from Yunnan province,the DNA probe results corre-lated well with blood smear ones.Of 107 P.falciparum samples determined by microscope ex-amination,99 were positive by sulfoprobe (92.5%);Of 72 P.vivax samples,1 was crosslyreacted;no cross reactions were found with any of 48 normal blood samples.It is indicated thatsulfoprobe may be useful in specific diagnosis and epidemiological investigation of P.falciparuminfection.

Field study of a soft X-ray aerosol neutralizer combined with electrostatic classifiers for nanoparticle size distribution measurements

Most conventional aerosol neutralizers are based on radioactive sources, which are controlled by strict regulations restricting their handling, transport, and storage. The TSI 3087 soft X-ray （SXR） neutralizer circumvents these legal restrictions. The aim of the present work is to compare the performance of a standalone SXR aerosol neutralizer with that of conventional radioactive aerosol neutralizers based on 85Kr （TSI 3077） and 241Am （Grimm 5522） by performing field tests in a real environmental scenario. The results obtained when the SXR neutralizer was connected to a mobility particle sizer spectrometer （MPS）, different from the device suggested by the manufacturer, were comparable with those obtained with the use of radioactive aerosol neutralizers. In changing the neutralizer, the particle number concentrations, measured with the MPS connected to the SXR neutralizer, almost remained within the 10% uncertainty bounds for the particle size interval 10-300 nm, when diffusion losses inside the SXR tube were considered. Based on our comparisons, the SXR neutralizer can be regarded as a standalone instrument that could solve the problems associated with legal restrictions on radioactive neutralizers and fulfil the need for a portable instrument for different field test purposes.