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On each day of follow-up, tympanic temperature was measured by electric thermometer and a finger prick blood sample was used for haemoglobin (Hb) measurement using a Hemocue photometer (Angelholm, Sweden), a microscopic slide, a 50 µL-blood sample for real-time nucleic acid sequence-based amplification (QT-NASBA) and a filter paper sample.
Plasmid Hmo41_It corresponding to one individual copy of HAmo SINE and Genomic DNA of silver carp and bighead carp were prepared as standard and sample for Real-Time PCR, respectively.
However, the sufficiency of the weekly sampling for real time predictions using ANN models needs to be further evaluated against longer weekly data sets as they become available.
All RNA samples for real time PCR were taken from the same pool of material as used for microarray analysis.
We recruited current and never smoker volunteers from Boston Medical Center for buccal (n = 11) and nasal (n = 15) microarray studies, and subsequent buccal epithelial samples for real competitive PCR validation using mass spectrometry (n = 14).
Biopsy samples for real-time (RT) analysis were directly transferred into RNAlater (QIAGEN) and stored at −80°C until further processing.
Plasmid Hmo41_It and Genomic DNA were prepared as the standard and samples for Real-Time PCR, respectively (Table 2).
The same was also done with the cycle 4 samples, resulting in a total of six samples for real-time qRT-PCR.
We used archived slides from these patients because we wanted to have the same blood samples for real-time RT-PCR and immunocytochemistry.
The copy numbers of the standard template were determined to quantitate PAR-2 mRNA level in samples for real-time reverse transcription (RT -PCR.
RNA sampling for real-time detection PCR was done in control snails and animals exposed to metals over a 5 day period (see above).
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