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In addition to on-chip LAMP detection, different LAMP products can be collected separately for further confirmatory tests using conventional gel electrophoresis and quantitative analysis, as demonstrated in Supporting Information Figures S-1 and 6 in laboratory settings, respectively.
LAMP detection and DNA preparation: The LAMP primers (Integrated DNA Technologies, Coralville, IA) for the target ctrA gene sequence from Neisseria meningitidis are shown in Supporting Information Table S-1.
As sensitivity was concerned, in comparison with regular PCR (ranging from 10-10 CFU/reaction) [ 6] and previous LAMP detection methodoloy (ranging from 10-10 cofies of DNA) [ 42], the detection limit of orfX-LAMP was found to be 10 copies DNA/tube and 10 CFU/reaction.
Fig. 1 LAMP detection of the ATCC 49926 porA gene using an automatic PCR cycler Fig. 2 Specificity of LAMP for porA detection.
The real-time fluorescence reverse transcription LAMP detection process can be completed within 35 min.
LAMP detection technology has the following advantages compared with other methods.
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The visual LAMP fluorescence detection is a simple method for rapid pathogen detection, from which a yes or no qualitative answer for POC detection can be achieved quickly based on the fluorescence of the LAMP products.
For the sensitivity of LAMP, the detection limit was 8.5 pg/μl (approximately 17 pg) DNA.
The advantage of real time-LAMP using SYTO 16 over end-point LAMP product detection is discussed.
Taking into account the advantages of LAMP, direct detection of MDV DNA in poultry dust has been conducted in this research.
Thus, the LAMP-TRBIV detection protocol has great potential for use in the detection of TRBIV in both the laboratory and the farm.
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